recombinant mouse il 23  (R&D Systems)

 
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    Name:
    Recombinant Mouse IL 23 Protein
    Description:
    The Recombinant Mouse IL 23 Protein from R D Systems is derived from Sf 21 stably transfected The Recombinant Mouse IL 23 Protein has been validated for the following applications Bioactivity
    Catalog Number:
    1887-ML-010
    Price:
    329
    Category:
    Proteins and Enzymes
    Source:
    Sf 21 (stably transfected)-derived Recombinant Mouse IL-23 Protein
    Applications:
    Bioactivity
    Purity:
    >90%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie« Blue Staining.
    Conjugate:
    Unconjugated
    Size:
    10 ug
    Buy from Supplier


    Structured Review

    R&D Systems recombinant mouse il 23
    Tnip1 controls <t>IL-23-mediated</t> skin disease. ( A ) qPCR analysis of mRNA levels in the ears of Tnip1 +/+ and Tnip1 −/− mice at day 14 during IL-23 injections ( n = 7 mice per group). ( B ) qPCR analysis of mRNA levels in the ears of Tnip1 −/−
    The Recombinant Mouse IL 23 Protein from R D Systems is derived from Sf 21 stably transfected The Recombinant Mouse IL 23 Protein has been validated for the following applications Bioactivity
    https://www.bioz.com/result/recombinant mouse il 23/product/R&D Systems
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    recombinant mouse il 23 - by Bioz Stars, 2021-05
    93/100 stars

    Images

    1) Product Images from "Keratinocytes contribute intrinsically to psoriasis upon loss of Tnip1 function"

    Article Title: Keratinocytes contribute intrinsically to psoriasis upon loss of Tnip1 function

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.1606996113

    Tnip1 controls IL-23-mediated skin disease. ( A ) qPCR analysis of mRNA levels in the ears of Tnip1 +/+ and Tnip1 −/− mice at day 14 during IL-23 injections ( n = 7 mice per group). ( B ) qPCR analysis of mRNA levels in the ears of Tnip1 −/−
    Figure Legend Snippet: Tnip1 controls IL-23-mediated skin disease. ( A ) qPCR analysis of mRNA levels in the ears of Tnip1 +/+ and Tnip1 −/− mice at day 14 during IL-23 injections ( n = 7 mice per group). ( B ) qPCR analysis of mRNA levels in the ears of Tnip1 −/−

    Techniques Used: Real-time Polymerase Chain Reaction, Mouse Assay

    Tnip1 controls IL-23–mediated skin disease. ( A–D ) The ears of Tnip1 +/+ and Tnip1 −/− mice ( n = 7 mice per group) were injected with IL-23 for 14 d and analyzed by measurement of ear thickness ( A ), microscopy of H E-stained
    Figure Legend Snippet: Tnip1 controls IL-23–mediated skin disease. ( A–D ) The ears of Tnip1 +/+ and Tnip1 −/− mice ( n = 7 mice per group) were injected with IL-23 for 14 d and analyzed by measurement of ear thickness ( A ), microscopy of H E-stained

    Techniques Used: Mouse Assay, Injection, Microscopy, Staining

    2) Product Images from "Inflammation, Immunity, Fibrosis, and Infection: IL-23 mediates murine liver transplantation ischemia-reperfusion injury via IFN-γ/IRF-1 pathway"

    Article Title: Inflammation, Immunity, Fibrosis, and Infection: IL-23 mediates murine liver transplantation ischemia-reperfusion injury via IFN-γ/IRF-1 pathway

    Journal: American Journal of Physiology - Gastrointestinal and Liver Physiology

    doi: 10.1152/ajpgi.00231.2018

    Blocking IL-23 with p19 antibody prevents IL-17, CXCL2, and IRF-1 expression in a mouse hepatocyte and nonparenchymal cell (HC/NPC) coculture system following hypoxia in vitro. A – C : IL-23 p19, IL-17, and chemokine (C-X-C motif) ligand 2 (CXCL2)
    Figure Legend Snippet: Blocking IL-23 with p19 antibody prevents IL-17, CXCL2, and IRF-1 expression in a mouse hepatocyte and nonparenchymal cell (HC/NPC) coculture system following hypoxia in vitro. A – C : IL-23 p19, IL-17, and chemokine (C-X-C motif) ligand 2 (CXCL2)

    Techniques Used: Blocking Assay, Expressing, In Vitro

    IL-17, chemokine (C-X-C motif) ligand 2 (CXCL2), IL-12, and IFN-γ are decreased by treatment with IL-23 antibody during cold ischemia-reperfusion (I/R) injury in mice. A – D : IL-17, CXCL2, IL-12, and IFN-γ mRNA expression was measured
    Figure Legend Snippet: IL-17, chemokine (C-X-C motif) ligand 2 (CXCL2), IL-12, and IFN-γ are decreased by treatment with IL-23 antibody during cold ischemia-reperfusion (I/R) injury in mice. A – D : IL-17, CXCL2, IL-12, and IFN-γ mRNA expression was measured

    Techniques Used: Mouse Assay, Expressing

    IL-23 antibody decreases ischemia-reperfusion (I/R) injury in wild-type (WT) but not natural killer T (NKT) cell-deficient mice. A : alanine aminotransferase (ALT) serum levels at 3 h following warm I/R with either IL-23 neutralizing antibody (2 μg/g
    Figure Legend Snippet: IL-23 antibody decreases ischemia-reperfusion (I/R) injury in wild-type (WT) but not natural killer T (NKT) cell-deficient mice. A : alanine aminotransferase (ALT) serum levels at 3 h following warm I/R with either IL-23 neutralizing antibody (2 μg/g

    Techniques Used: Mouse Assay

    IL-23 is elevated in vitro following hypoxia in a mouse hepatocyte and nonparenchymal cell (HC/NPC) coculture system. A : IL-23 mRNA levels as measured by p19 and p40 subunits in HC/NPC coculture system following hypoxia (1% O 2 ). B : Western blot showing
    Figure Legend Snippet: IL-23 is elevated in vitro following hypoxia in a mouse hepatocyte and nonparenchymal cell (HC/NPC) coculture system. A : IL-23 mRNA levels as measured by p19 and p40 subunits in HC/NPC coculture system following hypoxia (1% O 2 ). B : Western blot showing

    Techniques Used: In Vitro, Western Blot

    Overexpression of IL-23 in mice through adenovirus vector leads to induction of IL-17, interferon regulatory factor-1 (IRF-1), polymorphonuclear neutrophil (PMN) recruitment, and apoptosis. A : Western blot for IL-17 protein expression in the liver following
    Figure Legend Snippet: Overexpression of IL-23 in mice through adenovirus vector leads to induction of IL-17, interferon regulatory factor-1 (IRF-1), polymorphonuclear neutrophil (PMN) recruitment, and apoptosis. A : Western blot for IL-17 protein expression in the liver following

    Techniques Used: Over Expression, Mouse Assay, Plasmid Preparation, Western Blot, Expressing

    IL-23, interferon regulatory factor-1 (IRF-1), IL-17, and chemokine (C-X-C motif) ligand 2 (CXCL2) are elevated during cold ischemia-reperfusion (I/R) injury in mice. A : quantitative RT-PCR (qRT-PCR) was used to determine mRNA levels of IL-23 in liver
    Figure Legend Snippet: IL-23, interferon regulatory factor-1 (IRF-1), IL-17, and chemokine (C-X-C motif) ligand 2 (CXCL2) are elevated during cold ischemia-reperfusion (I/R) injury in mice. A : quantitative RT-PCR (qRT-PCR) was used to determine mRNA levels of IL-23 in liver

    Techniques Used: Mouse Assay, Quantitative RT-PCR

    Liver damage is attenuated during cold ischemia-reperfusion (I/R) injury by treatment with anti-IL-23 antibody. A and B : aspartate aminotransferase (AST)/alanine aminotransferase (ALT) serum levels at 6 h and 24 h following cold I/R with 18 h cold storage
    Figure Legend Snippet: Liver damage is attenuated during cold ischemia-reperfusion (I/R) injury by treatment with anti-IL-23 antibody. A and B : aspartate aminotransferase (AST)/alanine aminotransferase (ALT) serum levels at 6 h and 24 h following cold I/R with 18 h cold storage

    Techniques Used: AST Assay

    3) Product Images from "A cytokine network involving IL-36γ, IL-23, and IL-22 promotes antimicrobial defense and recovery from intestinal barrier damage"

    Article Title: A cytokine network involving IL-36γ, IL-23, and IL-22 promotes antimicrobial defense and recovery from intestinal barrier damage

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.1718902115

    Systemic IL-23 administration induces IL-22 and AMPs and rescues Il1rl2 −/− mice from DSS-induced colonic damage. ( A ) IL-22 protein expression in colons from Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS for 5 d in the presence or absence of IL-23. S100A8 ( B ), S100A9 ( C ), Reg3α ( D ), Reg3β ( E ), and Reg3γ ( F ) mRNA expression is shown in colons isolated from mice as in A . Data are representative of two independent experiments with five to six mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: ** P
    Figure Legend Snippet: Systemic IL-23 administration induces IL-22 and AMPs and rescues Il1rl2 −/− mice from DSS-induced colonic damage. ( A ) IL-22 protein expression in colons from Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS for 5 d in the presence or absence of IL-23. S100A8 ( B ), S100A9 ( C ), Reg3α ( D ), Reg3β ( E ), and Reg3γ ( F ) mRNA expression is shown in colons isolated from mice as in A . Data are representative of two independent experiments with five to six mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: ** P

    Techniques Used: Mouse Assay, Expressing, Isolation

    Systemic IL-23 administration induces resolution of DSS-induced colonic damage in Il1rl2 −/− mice. ( A ) DAI of Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS for 5 d, followed by normal water for 7 d, in the presence or absence of IL-23. ( B and C ) Image and length of colons from mice treated as in A . (Scale bar, 1 cm.) The H E staining ( D ) and histology scoring ( E ) of colon sections from mice treated as in A are shown. (Scale bar, 100 μm.) Data are representative of three independent experiments with four to five mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: * P
    Figure Legend Snippet: Systemic IL-23 administration induces resolution of DSS-induced colonic damage in Il1rl2 −/− mice. ( A ) DAI of Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS for 5 d, followed by normal water for 7 d, in the presence or absence of IL-23. ( B and C ) Image and length of colons from mice treated as in A . (Scale bar, 1 cm.) The H E staining ( D ) and histology scoring ( E ) of colon sections from mice treated as in A are shown. (Scale bar, 100 μm.) Data are representative of three independent experiments with four to five mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: * P

    Techniques Used: Mouse Assay, Staining

    IL-36γ–induced IL-22 production in colonic explants from DSS-treated mice is IL-23–dependent. ( A and B ) Colonic explants from control (no DSS) or 3-d DSS-treated (3 d DSS) WT mice were cultured for 60 h in the presence or absence of IL-36γ. Supernatants were analyzed for IL-23 ( A ) and IL-22 ( B ) by ELISA. ( C ) Colonic explants from 3-d DSS-treated WT mice were stimulated with IL-36γ and αp19 or αp40 antibodies for 60 h, and IL-22 expression was assessed by ELISA. ( D ) Colonic explants from 3-d DSS-treated WT ( Il12b +/+ ) or Il12b −/− mice stimulated with IL-36γ or IL-23 for 60 h and IL-22 expression were assessed by ELISA. Data are representative of two independent experiments with four to five mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: ** P
    Figure Legend Snippet: IL-36γ–induced IL-22 production in colonic explants from DSS-treated mice is IL-23–dependent. ( A and B ) Colonic explants from control (no DSS) or 3-d DSS-treated (3 d DSS) WT mice were cultured for 60 h in the presence or absence of IL-36γ. Supernatants were analyzed for IL-23 ( A ) and IL-22 ( B ) by ELISA. ( C ) Colonic explants from 3-d DSS-treated WT mice were stimulated with IL-36γ and αp19 or αp40 antibodies for 60 h, and IL-22 expression was assessed by ELISA. ( D ) Colonic explants from 3-d DSS-treated WT ( Il12b +/+ ) or Il12b −/− mice stimulated with IL-36γ or IL-23 for 60 h and IL-22 expression were assessed by ELISA. Data are representative of two independent experiments with four to five mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: ** P

    Techniques Used: Mouse Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Expressing

    IL-36R deficiency results in impaired IL-23 and IL-22 expression in the colons of DSS-treated mice. ( A ) PCR array gene expression analyses from colon tissues of Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS for 5 d. The time course of IL-23 mRNA ( B ) and protein ( C ) expression in colons from Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS is shown. The time course of IL-22 mRNA ( D ) and protein ( E ) expression in colons from Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS is shown. Data are representative of three independent experiments with three to four mice per group. All data are presented as mean ± SEM. * P
    Figure Legend Snippet: IL-36R deficiency results in impaired IL-23 and IL-22 expression in the colons of DSS-treated mice. ( A ) PCR array gene expression analyses from colon tissues of Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS for 5 d. The time course of IL-23 mRNA ( B ) and protein ( C ) expression in colons from Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS is shown. The time course of IL-22 mRNA ( D ) and protein ( E ) expression in colons from Il1rl2 +/+ and Il1rl2 −/− mice treated with DSS is shown. Data are representative of three independent experiments with three to four mice per group. All data are presented as mean ± SEM. * P

    Techniques Used: Expressing, Mouse Assay, Polymerase Chain Reaction

    IL-36γ induces IL-23 via signaling through c-Rel and NF-κBp50. ( A ) BMDCs were generated from c-rel +/+ and c-rel −/− mice and cultured in the presence or absence of IL-36γ for 24 h, and IL-23 was assessed by ELISA. ( B and C ) WT BMDCs were cultured in the presence or absence of IL-36γ for 24 h, and IL-23 was assessed by ELISA. ( B ) Some cultures were pretreated with the c-Rel inhibitor (IT-603) or with vehicle control (DMSO) for 1 h. ( C ) Some cultures were pretreated with p50 or p65 inhibitor peptides or control peptides for 1 h. ( D ) BMDCs were generated from p50 +/+ and p50 −/− mice and cultured in the presence or absence of IL-36γ for 24 h, and IL-23 was assessed by ELISA. ( E ) ChIP assays for p50 and c-Rel binding to the p19 promoter in BMDCs treated with IL-36γ for 8 h. Data in A – D are representative of at least two independent experiments with n = 5 mice. Data in E are the combined data of two independent experiments with three replicates per experiment. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: ** P
    Figure Legend Snippet: IL-36γ induces IL-23 via signaling through c-Rel and NF-κBp50. ( A ) BMDCs were generated from c-rel +/+ and c-rel −/− mice and cultured in the presence or absence of IL-36γ for 24 h, and IL-23 was assessed by ELISA. ( B and C ) WT BMDCs were cultured in the presence or absence of IL-36γ for 24 h, and IL-23 was assessed by ELISA. ( B ) Some cultures were pretreated with the c-Rel inhibitor (IT-603) or with vehicle control (DMSO) for 1 h. ( C ) Some cultures were pretreated with p50 or p65 inhibitor peptides or control peptides for 1 h. ( D ) BMDCs were generated from p50 +/+ and p50 −/− mice and cultured in the presence or absence of IL-36γ for 24 h, and IL-23 was assessed by ELISA. ( E ) ChIP assays for p50 and c-Rel binding to the p19 promoter in BMDCs treated with IL-36γ for 8 h. Data in A – D are representative of at least two independent experiments with n = 5 mice. Data in E are the combined data of two independent experiments with three replicates per experiment. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: ** P

    Techniques Used: Generated, Mouse Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Chromatin Immunoprecipitation, Binding Assay

    Notch2-dependent DCs are required for IL-36γ–induced IL-23 and IL-22 expression and recovery from colonic damage. ( A ) IL-36R ( Il1rl2 ) mRNA expression was analyzed by qPCR in colon tissue isolated from DSS-treated batf3 +/+ , batf3 −/− , Notch2 fl/fl , and Notch2 cKO mice directly ex vivo. ( B and C ) Colonic explants from DSS-treated mice were cultured for 60 h in the presence or absence of IL-36γ or IL-23. Supernatants were analyzed for IL-23 ( B ) and IL-22 ( C ) expression by ELISA. ( D ) DAI of batf3 +/+ , batf3 −/− , Notch2 fl/fl , and Notch2 cKO mice treated with DSS for 5 d, followed by normal water. ( E ) Image and colon length from mice treated as in D , at day 14. (Scale bar, 1 cm.) The expression of IL-23 ( F ) and IL-22 ( G ) in colon tissues from DSS mice at day 5 is shown. Data are representative of two independent experiments with three to four mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: * P
    Figure Legend Snippet: Notch2-dependent DCs are required for IL-36γ–induced IL-23 and IL-22 expression and recovery from colonic damage. ( A ) IL-36R ( Il1rl2 ) mRNA expression was analyzed by qPCR in colon tissue isolated from DSS-treated batf3 +/+ , batf3 −/− , Notch2 fl/fl , and Notch2 cKO mice directly ex vivo. ( B and C ) Colonic explants from DSS-treated mice were cultured for 60 h in the presence or absence of IL-36γ or IL-23. Supernatants were analyzed for IL-23 ( B ) and IL-22 ( C ) expression by ELISA. ( D ) DAI of batf3 +/+ , batf3 −/− , Notch2 fl/fl , and Notch2 cKO mice treated with DSS for 5 d, followed by normal water. ( E ) Image and colon length from mice treated as in D , at day 14. (Scale bar, 1 cm.) The expression of IL-23 ( F ) and IL-22 ( G ) in colon tissues from DSS mice at day 5 is shown. Data are representative of two independent experiments with three to four mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test: * P

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction, Isolation, Mouse Assay, Ex Vivo, Cell Culture, Enzyme-linked Immunosorbent Assay

    4) Product Images from "CCR6 is required for epidermal trafficking of ?? T cells in an IL-23-induced model of psoriasiform dermatitis"

    Article Title: CCR6 is required for epidermal trafficking of ?? T cells in an IL-23-induced model of psoriasiform dermatitis

    Journal: The Journal of investigative dermatology

    doi: 10.1038/jid.2012.260

    Neutralizing anti-CCL20 monoclonal antibodies inhibit the trafficking of GDL T cells in response to IL-23 injection Following treatment with neutralizing anti-CCL20 mAb, isotype control, or PBS injection for 5 days, both pooled epidermal ( a ) and dermal ( b ) cell suspensions from 4 mouse ears (per condition) were stained with mAbs against γδ TCR for flow cytometry e( a,b ) or processed for RNA extraction for RT-PCR ( c,d ). In ( a,b ), the percentage indicated in each dot plot reflects the number of GDL cells (i.e., events) in the oval window divided by the total sample events shown in the entire dot plot. Percent reduction in epidermal GDL T cells was calculated at 76% for data shown [(% GDL cells IL23 + isotype)-(% GDL cells IL23+anti-CCL20)]/[(%GDL cells IL23+isotype)-(%GDL cells PBS)]*100. Pooled epidermal ( c ) and dermal ( d ) cell suspensions (see above) were used for quantitative RT-PCR to measure expression of IL-22 and CCR6. Fold changes were calculated for IL-22 and Ccr6 mRNAs normalized for Gapdh mRNA vs . the PBS treated sample. Error bars indicate SD ( c,d ) based on a single, pooled treatment group that was divided into three wells for RT-PCR measurement. Similar results were obtained in 2 independent experiments.
    Figure Legend Snippet: Neutralizing anti-CCL20 monoclonal antibodies inhibit the trafficking of GDL T cells in response to IL-23 injection Following treatment with neutralizing anti-CCL20 mAb, isotype control, or PBS injection for 5 days, both pooled epidermal ( a ) and dermal ( b ) cell suspensions from 4 mouse ears (per condition) were stained with mAbs against γδ TCR for flow cytometry e( a,b ) or processed for RNA extraction for RT-PCR ( c,d ). In ( a,b ), the percentage indicated in each dot plot reflects the number of GDL cells (i.e., events) in the oval window divided by the total sample events shown in the entire dot plot. Percent reduction in epidermal GDL T cells was calculated at 76% for data shown [(% GDL cells IL23 + isotype)-(% GDL cells IL23+anti-CCL20)]/[(%GDL cells IL23+isotype)-(%GDL cells PBS)]*100. Pooled epidermal ( c ) and dermal ( d ) cell suspensions (see above) were used for quantitative RT-PCR to measure expression of IL-22 and CCR6. Fold changes were calculated for IL-22 and Ccr6 mRNAs normalized for Gapdh mRNA vs . the PBS treated sample. Error bars indicate SD ( c,d ) based on a single, pooled treatment group that was divided into three wells for RT-PCR measurement. Similar results were obtained in 2 independent experiments.

    Techniques Used: Injection, Staining, Flow Cytometry, Cytometry, RNA Extraction, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Expressing

    Neutralizing anti-CCL20 monoclonal antibodies inhibit the ear swelling in response to IL-23 injection ( a ) WT mice were injected with neutralizing anti-CCL20 mAb, isotype control, or PBS in ear skin with subsequent measurement of ear thickness throughout the treatment period. *P
    Figure Legend Snippet: Neutralizing anti-CCL20 monoclonal antibodies inhibit the ear swelling in response to IL-23 injection ( a ) WT mice were injected with neutralizing anti-CCL20 mAb, isotype control, or PBS in ear skin with subsequent measurement of ear thickness throughout the treatment period. *P

    Techniques Used: Injection, Mouse Assay

    CCL20 expression in mouse epidermis is upregulated within 24 hr of IL-23 injection IL-23 was injected into ears of WT mice at day 0, 2, and 4. Mice were euthanized on the indicated days and ears were stained with anti-CCL20 antibody (red) and DAPI (nuclear counterstain in blue). Isotype control staining showed no epidermal staining with either IL-23 or PBS-injected ears (data not shown). Scale bar = 50 μm.
    Figure Legend Snippet: CCL20 expression in mouse epidermis is upregulated within 24 hr of IL-23 injection IL-23 was injected into ears of WT mice at day 0, 2, and 4. Mice were euthanized on the indicated days and ears were stained with anti-CCL20 antibody (red) and DAPI (nuclear counterstain in blue). Isotype control staining showed no epidermal staining with either IL-23 or PBS-injected ears (data not shown). Scale bar = 50 μm.

    Techniques Used: Expressing, Injection, Mouse Assay, Staining

    Immunofluorescence staining for CCR6 expression following IL-23 injection Following IL-23 ( a ) or PBS ( b ) injection for 6 days, mice were euthanized and ears were stained with anti-CCR6 mAbs (green) and anti-Laminin 332 mAbs (basement membrane in red), and the 3D images were acquired using a confocal microscope. The figure shows the perspective from the dermis toward the epidermis. The width of basement membrane in red shows the depth of the sections. Larger numbers of CCR6-expressing cells in the epidermis (above the basement membrane) were revealed in IL-23-, but not PBS-injected, mice. CCR6-expressing cells ( a ) at (arrow head), immediately adjacent to (double arrowheads), or above (arrow) the basement membrane in the dashed rectangular area marked in the upper figure are shown at higher magnification below. Isotype control staining showed no staining with either IL-23 or PBS-injected ears (data not shown). Scale bar = 10 μm. In ( c ), IL-23-treated skin sections were stained with labeled mAbs against CCR6 (green) and the γδ-TCR (red). Images were acquired by confocal microscopy at the level of the epidermis (x63 objective, scale bar as indicated). Similar results were obtained in 3 independent experiments.
    Figure Legend Snippet: Immunofluorescence staining for CCR6 expression following IL-23 injection Following IL-23 ( a ) or PBS ( b ) injection for 6 days, mice were euthanized and ears were stained with anti-CCR6 mAbs (green) and anti-Laminin 332 mAbs (basement membrane in red), and the 3D images were acquired using a confocal microscope. The figure shows the perspective from the dermis toward the epidermis. The width of basement membrane in red shows the depth of the sections. Larger numbers of CCR6-expressing cells in the epidermis (above the basement membrane) were revealed in IL-23-, but not PBS-injected, mice. CCR6-expressing cells ( a ) at (arrow head), immediately adjacent to (double arrowheads), or above (arrow) the basement membrane in the dashed rectangular area marked in the upper figure are shown at higher magnification below. Isotype control staining showed no staining with either IL-23 or PBS-injected ears (data not shown). Scale bar = 10 μm. In ( c ), IL-23-treated skin sections were stained with labeled mAbs against CCR6 (green) and the γδ-TCR (red). Images were acquired by confocal microscopy at the level of the epidermis (x63 objective, scale bar as indicated). Similar results were obtained in 3 independent experiments.

    Techniques Used: Immunofluorescence, Staining, Expressing, Injection, Mouse Assay, Microscopy, Labeling, Confocal Microscopy

    CCR6 KO mice fail to accumulate IL-22 + , GDL T cells in response to IL-23 injection Following IL-23 injection for 6 days, both WT and CCR6 KO mice were euthanized. Both epidermal and dermal cell suspensions were analyzed by flow cytometry with γδ TCR, CD3, and IL-22 mAbs. ( a ) Total numbers of GDL T cells per PBS- or IL-23-injected ear of WT and CCR6 KO mice were counted. The percentages of CD3 + GDL T cells ( b ) and IL-22 + GDL T cells ( c ) were calculated. *P
    Figure Legend Snippet: CCR6 KO mice fail to accumulate IL-22 + , GDL T cells in response to IL-23 injection Following IL-23 injection for 6 days, both WT and CCR6 KO mice were euthanized. Both epidermal and dermal cell suspensions were analyzed by flow cytometry with γδ TCR, CD3, and IL-22 mAbs. ( a ) Total numbers of GDL T cells per PBS- or IL-23-injected ear of WT and CCR6 KO mice were counted. The percentages of CD3 + GDL T cells ( b ) and IL-22 + GDL T cells ( c ) were calculated. *P

    Techniques Used: Mouse Assay, Injection, Flow Cytometry, Cytometry

    5) Product Images from "RIG‐I antiviral signaling drives interleukin‐23 production and psoriasis‐like skin disease"

    Article Title: RIG‐I antiviral signaling drives interleukin‐23 production and psoriasis‐like skin disease

    Journal: EMBO Molecular Medicine

    doi: 10.15252/emmm.201607027

    NF‐κB signaling is required for the expression of IL‐23p19 in DCs BMDCs were transfected with scramble siRNA (Ctr), p65 siRNA (si‐p65), IRF‐3 siRNA (si‐IRF‐3), or IRF‐7 siRNA (si‐IRF‐7). Cell lysates were immunoblotted with anti‐p65, anti‐IRF‐3, anti‐IRF‐7, or anti‐β‐actin Ab. Values were expressed as fold changes relative to controls and normalized to β‐actin. qPCR and ELISA analysis of IL‐23p19 expression in BMDCs transfected with scramble siRNA (Ctr), p65 siRNA (si‐p65) (B), IRF‐3 siRNA (si‐IRF‐3) (C), or IRF‐7 siRNA (si‐IRF‐7) (D), and stimulated with IL‐23 or 5′ppp‐dsRNA for 30 or 36 h. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–6 per group (mean ± SEM). The schematic diagram shows one potential binding site of p65 in the putative promoter element of mouse IL‐23p19. p65 was immunoprecipitated from BMDCs stimulated with IL‐23. Immunoprecipitates were assayed for the enrichment of IL‐23 promoter. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–4 per group (mean ± SEM). Luciferase activity in lysates of RAW264.7 cells transfected with luciferase reporter plasmids of pGL3‐basic empty vector (basic), IL‐23p19 promoter (pIL‐23p19‐wt), or IL‐23p19 promoter with mutation on the predicted NF‐κB binding site (pIL‐23p19‐mut), unstimulated or stimulated with IL‐23 or/and 5′ppp‐dsRNA. Results are presented as the ratio of firefly luciferase to Renilla luciferase activity, relative to that of unstimulated RAW264.7 cells transfected with pGL3‐basic empty vector. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–4 per group (mean ± SEM). Source data are available online for this figure.
    Figure Legend Snippet: NF‐κB signaling is required for the expression of IL‐23p19 in DCs BMDCs were transfected with scramble siRNA (Ctr), p65 siRNA (si‐p65), IRF‐3 siRNA (si‐IRF‐3), or IRF‐7 siRNA (si‐IRF‐7). Cell lysates were immunoblotted with anti‐p65, anti‐IRF‐3, anti‐IRF‐7, or anti‐β‐actin Ab. Values were expressed as fold changes relative to controls and normalized to β‐actin. qPCR and ELISA analysis of IL‐23p19 expression in BMDCs transfected with scramble siRNA (Ctr), p65 siRNA (si‐p65) (B), IRF‐3 siRNA (si‐IRF‐3) (C), or IRF‐7 siRNA (si‐IRF‐7) (D), and stimulated with IL‐23 or 5′ppp‐dsRNA for 30 or 36 h. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–6 per group (mean ± SEM). The schematic diagram shows one potential binding site of p65 in the putative promoter element of mouse IL‐23p19. p65 was immunoprecipitated from BMDCs stimulated with IL‐23. Immunoprecipitates were assayed for the enrichment of IL‐23 promoter. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–4 per group (mean ± SEM). Luciferase activity in lysates of RAW264.7 cells transfected with luciferase reporter plasmids of pGL3‐basic empty vector (basic), IL‐23p19 promoter (pIL‐23p19‐wt), or IL‐23p19 promoter with mutation on the predicted NF‐κB binding site (pIL‐23p19‐mut), unstimulated or stimulated with IL‐23 or/and 5′ppp‐dsRNA. Results are presented as the ratio of firefly luciferase to Renilla luciferase activity, relative to that of unstimulated RAW264.7 cells transfected with pGL3‐basic empty vector. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–4 per group (mean ± SEM). Source data are available online for this figure.

    Techniques Used: Expressing, Transfection, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Binding Assay, Immunoprecipitation, Luciferase, Activity Assay, Plasmid Preparation, Mutagenesis

    Germ‐free mice are almost protected from IL‐23‐induced psoriasis‐like skin inflammation The ear thickness of WT mice in specific pathogen‐free (SPF) conditions or in germ‐free (GF) conditions treated with PBS or IL‐23. Data are presented on the indicated day relative to day 0. Significant differences are indicated: P
    Figure Legend Snippet: Germ‐free mice are almost protected from IL‐23‐induced psoriasis‐like skin inflammation The ear thickness of WT mice in specific pathogen‐free (SPF) conditions or in germ‐free (GF) conditions treated with PBS or IL‐23. Data are presented on the indicated day relative to day 0. Significant differences are indicated: P

    Techniques Used: Mouse Assay

    Schematic diagram of how the antiviral signaling mediates psoriasis pathogenesis In genetically predisposed individuals, the virus infection causes the activation of TLR‐7/8 and/or RIG‐I, and subsequently triggers IL‐23 release by CD11c + DCs via the NF‐κB pathway. Genetic mutations in NF‐κB‐related genes result in an impaired negative regulation of its proinflammatory activity accompanied by uncontrolled IL‐23 release, thus leading to psoriasis.
    Figure Legend Snippet: Schematic diagram of how the antiviral signaling mediates psoriasis pathogenesis In genetically predisposed individuals, the virus infection causes the activation of TLR‐7/8 and/or RIG‐I, and subsequently triggers IL‐23 release by CD11c + DCs via the NF‐κB pathway. Genetic mutations in NF‐κB‐related genes result in an impaired negative regulation of its proinflammatory activity accompanied by uncontrolled IL‐23 release, thus leading to psoriasis.

    Techniques Used: Infection, Activation Assay, Activity Assay

    RIG‐I expression in non‐haematopoietic cells is not required for IL‐23‐induced psoriasis‐like skin inflammation Lethally irradiated WT and RIG‐I −/− mice were adoptively transferred with WT bone marrow (BM) cells, and the generated chimeric mice were subjected to IL‐23‐induced psoriasis‐like skin inflammation. Data are presented on the indicated day relative to day 0. Significant differences are indicated: one‐way ANOVA, n = 5 per group (mean ± SEM). Representative H E staining and Ki67 immunostaining of the ears treated as in (A), n = 5 per group. Scale bar: 50 μm. Acanthosis (C) and dermal cellular infiltrates (D) of WT BM‐WT or WT BM‐RIG‐I −/− mice treated with IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 5 per group (mean ± SEM).
    Figure Legend Snippet: RIG‐I expression in non‐haematopoietic cells is not required for IL‐23‐induced psoriasis‐like skin inflammation Lethally irradiated WT and RIG‐I −/− mice were adoptively transferred with WT bone marrow (BM) cells, and the generated chimeric mice were subjected to IL‐23‐induced psoriasis‐like skin inflammation. Data are presented on the indicated day relative to day 0. Significant differences are indicated: one‐way ANOVA, n = 5 per group (mean ± SEM). Representative H E staining and Ki67 immunostaining of the ears treated as in (A), n = 5 per group. Scale bar: 50 μm. Acanthosis (C) and dermal cellular infiltrates (D) of WT BM‐WT or WT BM‐RIG‐I −/− mice treated with IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 5 per group (mean ± SEM).

    Techniques Used: Expressing, Irradiation, Mouse Assay, Generated, Staining, Immunostaining, Two Tailed Test

    RIG‐I expression in DCs is critical for IL‐23 production Representative flow cytometry analysis of the frequency of CD45 + RIG‐I + cells in WT ears treated with PBS or IL‐23 ( n = 5). Statistical analysis of the results in (A). Significant differences are indicated: two‐tailed Student's t ‐test, n = 5 per group (mean ± SEM). Representative flow cytometry analysis of the frequency of RIG‐I + CD3 + , RIG‐I + CD11c + , and RIG‐I + F4/80 + cells in ears treated with IL‐23 ( n = 4–5). Statistical analysis of the results in (C). Significant differences are indicated: two‐tailed Student's t ‐test, n = 4–5 per group (mean ± SEM). qPCR analysis of IL‐23p19 mRNA expression of cultured WT BMDCs and RIG‐I −/− BMDCs treated with indicated stimulation for 24 h. qPCR values expressed as the ratio of mRNA to β‐actin, relative to negative control, and indicated as fold change. Significant differences are indicated: two‐tailed Student's t ‐test, n = 5–6 per group (mean ± SEM). ELISA detection of IL‐23p19 protein levels in supernatants of cultured WT BMDCs and RIG‐I −/− BMDCs treated with indicated stimulation for 48 h. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3 per group (mean ± SEM).
    Figure Legend Snippet: RIG‐I expression in DCs is critical for IL‐23 production Representative flow cytometry analysis of the frequency of CD45 + RIG‐I + cells in WT ears treated with PBS or IL‐23 ( n = 5). Statistical analysis of the results in (A). Significant differences are indicated: two‐tailed Student's t ‐test, n = 5 per group (mean ± SEM). Representative flow cytometry analysis of the frequency of RIG‐I + CD3 + , RIG‐I + CD11c + , and RIG‐I + F4/80 + cells in ears treated with IL‐23 ( n = 4–5). Statistical analysis of the results in (C). Significant differences are indicated: two‐tailed Student's t ‐test, n = 4–5 per group (mean ± SEM). qPCR analysis of IL‐23p19 mRNA expression of cultured WT BMDCs and RIG‐I −/− BMDCs treated with indicated stimulation for 24 h. qPCR values expressed as the ratio of mRNA to β‐actin, relative to negative control, and indicated as fold change. Significant differences are indicated: two‐tailed Student's t ‐test, n = 5–6 per group (mean ± SEM). ELISA detection of IL‐23p19 protein levels in supernatants of cultured WT BMDCs and RIG‐I −/− BMDCs treated with indicated stimulation for 48 h. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3 per group (mean ± SEM).

    Techniques Used: Expressing, Flow Cytometry, Cytometry, Two Tailed Test, Real-time Polymerase Chain Reaction, Cell Culture, Negative Control, Enzyme-linked Immunosorbent Assay

    Inflammatory cell infiltrates and IL‐17 expression levels in IL‐23‐induced mouse model Representative immunostaining of CD3 in ear skin derived from WT and RIG‐I −/− mice treated with PBS or IL‐23, n = 3–5 per group. Scale bar: 20 μm. Quantitation of CD3 + cells in ear skin derived from WT and RIG‐I −/− mice treated with PBS or IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–5 per group (mean ± SEM). Representative immunofluorescence staining of CD11c in ear skin derived from WT and RIG‐I −/− mice treated with IL‐23, n = 3–5 per group. Scale bar: 50 μm. ELISA detection of IL‐17 protein levels in supernatants of ear skin homogenates derived from indicated groups. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3 per group (mean ± SEM).
    Figure Legend Snippet: Inflammatory cell infiltrates and IL‐17 expression levels in IL‐23‐induced mouse model Representative immunostaining of CD3 in ear skin derived from WT and RIG‐I −/− mice treated with PBS or IL‐23, n = 3–5 per group. Scale bar: 20 μm. Quantitation of CD3 + cells in ear skin derived from WT and RIG‐I −/− mice treated with PBS or IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–5 per group (mean ± SEM). Representative immunofluorescence staining of CD11c in ear skin derived from WT and RIG‐I −/− mice treated with IL‐23, n = 3–5 per group. Scale bar: 50 μm. ELISA detection of IL‐17 protein levels in supernatants of ear skin homogenates derived from indicated groups. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3 per group (mean ± SEM).

    Techniques Used: Expressing, Immunostaining, Derivative Assay, Mouse Assay, Quantitation Assay, Two Tailed Test, Immunofluorescence, Staining, Enzyme-linked Immunosorbent Assay

    IL‐23‐induced psoriasis‐like skin disease is attenuated in RIG‐I −/− mice Representative photographs of the ears of WT mice (upper panel) and RIG‐I −/− mice (lower panel) after intradermal injection with PBS or IL‐23 (500 ng) on every other day for eight times, n = 4–5 per group. The ear thickness of WT and RIG‐I −/− mice on the indicated day presented relative to day 0. Significant differences are indicated: P = 0.0033, one‐way ANOVA, n = 4–5 per group (mean ± SEM). Representative H E staining of the ears treated as in (A), n = 4–5 per group. Scale bar: 50 μm. Acanthosis of WT and RIG‐I −/− mice treated with PBS or IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–8 per group (mean ± SEM). Representative immunostaining of Ki67 in ear skin derived from WT and RIG‐I −/− mice treated with PBS or IL‐23, n = 3–6 per group. Scale bar: 20 μm. Quantitation of Ki67 + cells in ear skin derived from WT and RIG‐I −/− mice treated with PBS or IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–6 per group (mean ± SEM). Western blot analysis of RIG‐I and p‐IκBα expression after treatment as in (A). ELISA detection of IL‐23p19 protein levels in supernatants of ear skin homogenates derived from indicated groups. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3 per group (mean ± SEM). Source data are available online for this figure.
    Figure Legend Snippet: IL‐23‐induced psoriasis‐like skin disease is attenuated in RIG‐I −/− mice Representative photographs of the ears of WT mice (upper panel) and RIG‐I −/− mice (lower panel) after intradermal injection with PBS or IL‐23 (500 ng) on every other day for eight times, n = 4–5 per group. The ear thickness of WT and RIG‐I −/− mice on the indicated day presented relative to day 0. Significant differences are indicated: P = 0.0033, one‐way ANOVA, n = 4–5 per group (mean ± SEM). Representative H E staining of the ears treated as in (A), n = 4–5 per group. Scale bar: 50 μm. Acanthosis of WT and RIG‐I −/− mice treated with PBS or IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–8 per group (mean ± SEM). Representative immunostaining of Ki67 in ear skin derived from WT and RIG‐I −/− mice treated with PBS or IL‐23, n = 3–6 per group. Scale bar: 20 μm. Quantitation of Ki67 + cells in ear skin derived from WT and RIG‐I −/− mice treated with PBS or IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–6 per group (mean ± SEM). Western blot analysis of RIG‐I and p‐IκBα expression after treatment as in (A). ELISA detection of IL‐23p19 protein levels in supernatants of ear skin homogenates derived from indicated groups. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3 per group (mean ± SEM). Source data are available online for this figure.

    Techniques Used: Mouse Assay, Injection, Staining, Two Tailed Test, Immunostaining, Derivative Assay, Quantitation Assay, Western Blot, Expressing, Enzyme-linked Immunosorbent Assay

    RIG‐I expression is increased in lesional skin of psoriasis mouse model and patients qPCR analysis of RIG‐I mRNA expression in psoriatic lesions compared to healthy samples. Significant differences are indicated: two‐tailed Student's t ‐test, n = 10–13 per group (mean ± SEM). Representative immunohistochemical detection of RIG‐I in skin of psoriatic lesions compared with healthy samples, n = 5 per group. Scale bar: 20 μm. Quantitation of RIG‐I + cells in dermis of psoriatic skin compared to healthy skin. Significant differences are indicated: two‐tailed Student's t ‐test, n = 5 per group (mean ± SEM). qPCR analysis of RIG‐I mRNA expression in ear skin of mice treated with PBS or IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 11 per group (mean ± SEM). Western blot analysis of RIG‐I expression in ears treated with PBS or IL‐23 ( n = 3). Western blot analysis of RIG‐I expression in ears untreated or treated with IMQ ( n = 4). Correlation between RIG‐I expression and acanthosis of 84 psoriatic skin samples ( r 2 = 0.3577). Significant differences are indicated: Pearson test, two‐tailed. Data information: qPCR values expressed as the ratio of mRNA to β‐actin, relative to control samples, and indicated as fold change. Source data are available online for this figure.
    Figure Legend Snippet: RIG‐I expression is increased in lesional skin of psoriasis mouse model and patients qPCR analysis of RIG‐I mRNA expression in psoriatic lesions compared to healthy samples. Significant differences are indicated: two‐tailed Student's t ‐test, n = 10–13 per group (mean ± SEM). Representative immunohistochemical detection of RIG‐I in skin of psoriatic lesions compared with healthy samples, n = 5 per group. Scale bar: 20 μm. Quantitation of RIG‐I + cells in dermis of psoriatic skin compared to healthy skin. Significant differences are indicated: two‐tailed Student's t ‐test, n = 5 per group (mean ± SEM). qPCR analysis of RIG‐I mRNA expression in ear skin of mice treated with PBS or IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 11 per group (mean ± SEM). Western blot analysis of RIG‐I expression in ears treated with PBS or IL‐23 ( n = 3). Western blot analysis of RIG‐I expression in ears untreated or treated with IMQ ( n = 4). Correlation between RIG‐I expression and acanthosis of 84 psoriatic skin samples ( r 2 = 0.3577). Significant differences are indicated: Pearson test, two‐tailed. Data information: qPCR values expressed as the ratio of mRNA to β‐actin, relative to control samples, and indicated as fold change. Source data are available online for this figure.

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction, Two Tailed Test, Immunohistochemistry, Quantitation Assay, Mouse Assay, Western Blot

    IL‐23‐induced psoriasis‐like skin disease is ameliorated in IL‐23 −/− mice Representative photographs of the ears of WT mice (left panel) and IL‐23 −/− mice (right panel) after intradermal injection with IL‐23 (500 ng) on every other day for 8 times, n = 6 per group. The ear thickness of WT and IL‐23 −/− mice on the indicated day presented relative to day 0. Significant differences are indicated: one‐way ANOVA, n = 6 per group (mean ± SEM). Representative H E staining of the ears treated as in (A), n = 6 per group. Scale bar: 50 μm. Acanthosis of WT and IL‐23 −/− mice treated with IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 6 per group (mean ± SEM). Representative immunostaining of Ki67 in ear skin derived from WT and IL‐23 −/− mice treated with IL‐23 n = 6 per group. Scale bar: 50 μm. Quantitation of Ki67 + cells in ear skin derived from WT and IL‐23 −/− mice treated with IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 6 per group (mean ± SEM). ELISA detection of IL‐23p19 (G) and IL‐17 (H) protein levels in supernatants of ear skin homogenates derived from indicated groups. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–5 per group (mean ± SEM).
    Figure Legend Snippet: IL‐23‐induced psoriasis‐like skin disease is ameliorated in IL‐23 −/− mice Representative photographs of the ears of WT mice (left panel) and IL‐23 −/− mice (right panel) after intradermal injection with IL‐23 (500 ng) on every other day for 8 times, n = 6 per group. The ear thickness of WT and IL‐23 −/− mice on the indicated day presented relative to day 0. Significant differences are indicated: one‐way ANOVA, n = 6 per group (mean ± SEM). Representative H E staining of the ears treated as in (A), n = 6 per group. Scale bar: 50 μm. Acanthosis of WT and IL‐23 −/− mice treated with IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 6 per group (mean ± SEM). Representative immunostaining of Ki67 in ear skin derived from WT and IL‐23 −/− mice treated with IL‐23 n = 6 per group. Scale bar: 50 μm. Quantitation of Ki67 + cells in ear skin derived from WT and IL‐23 −/− mice treated with IL‐23. Significant differences are indicated: two‐tailed Student's t ‐test, n = 6 per group (mean ± SEM). ELISA detection of IL‐23p19 (G) and IL‐17 (H) protein levels in supernatants of ear skin homogenates derived from indicated groups. Significant differences are indicated: two‐tailed Student's t ‐test, n = 3–5 per group (mean ± SEM).

    Techniques Used: Mouse Assay, Injection, Staining, Two Tailed Test, Immunostaining, Derivative Assay, Quantitation Assay, Enzyme-linked Immunosorbent Assay

    RIG‐I expression in haematopoietic cells is critical for IL‐23‐induced psoriasis‐like skin inflammation Lethally irradiated WT mice were adoptively transferred with WT or RIG‐I −/− bone marrow cells, and the generated chimeric mice were subjected to IL‐23‐induced psoriasis‐like skin inflammation. Graph represents the ear thickness of WT recipient mice on the indicated day relative to day 0. Significant differences are indicated: P
    Figure Legend Snippet: RIG‐I expression in haematopoietic cells is critical for IL‐23‐induced psoriasis‐like skin inflammation Lethally irradiated WT mice were adoptively transferred with WT or RIG‐I −/− bone marrow cells, and the generated chimeric mice were subjected to IL‐23‐induced psoriasis‐like skin inflammation. Graph represents the ear thickness of WT recipient mice on the indicated day relative to day 0. Significant differences are indicated: P

    Techniques Used: Expressing, Irradiation, Mouse Assay, Generated

    6) Product Images from "Inflammation, Immunity, Fibrosis, and Infection: IL-23 mediates murine liver transplantation ischemia-reperfusion injury via IFN-γ/IRF-1 pathway"

    Article Title: Inflammation, Immunity, Fibrosis, and Infection: IL-23 mediates murine liver transplantation ischemia-reperfusion injury via IFN-γ/IRF-1 pathway

    Journal: American Journal of Physiology - Gastrointestinal and Liver Physiology

    doi: 10.1152/ajpgi.00231.2018

    Blocking IL-23 with p19 antibody prevents IL-17, CXCL2, and IRF-1 expression in a mouse hepatocyte and nonparenchymal cell (HC/NPC) coculture system following hypoxia in vitro. A – C : IL-23 p19, IL-17, and chemokine (C-X-C motif) ligand 2 (CXCL2)
    Figure Legend Snippet: Blocking IL-23 with p19 antibody prevents IL-17, CXCL2, and IRF-1 expression in a mouse hepatocyte and nonparenchymal cell (HC/NPC) coculture system following hypoxia in vitro. A – C : IL-23 p19, IL-17, and chemokine (C-X-C motif) ligand 2 (CXCL2)

    Techniques Used: Blocking Assay, Expressing, In Vitro

    IL-17, chemokine (C-X-C motif) ligand 2 (CXCL2), IL-12, and IFN-γ are decreased by treatment with IL-23 antibody during cold ischemia-reperfusion (I/R) injury in mice. A – D : IL-17, CXCL2, IL-12, and IFN-γ mRNA expression was measured
    Figure Legend Snippet: IL-17, chemokine (C-X-C motif) ligand 2 (CXCL2), IL-12, and IFN-γ are decreased by treatment with IL-23 antibody during cold ischemia-reperfusion (I/R) injury in mice. A – D : IL-17, CXCL2, IL-12, and IFN-γ mRNA expression was measured

    Techniques Used: Mouse Assay, Expressing

    IL-23 antibody decreases ischemia-reperfusion (I/R) injury in wild-type (WT) but not natural killer T (NKT) cell-deficient mice. A : alanine aminotransferase (ALT) serum levels at 3 h following warm I/R with either IL-23 neutralizing antibody (2 μg/g
    Figure Legend Snippet: IL-23 antibody decreases ischemia-reperfusion (I/R) injury in wild-type (WT) but not natural killer T (NKT) cell-deficient mice. A : alanine aminotransferase (ALT) serum levels at 3 h following warm I/R with either IL-23 neutralizing antibody (2 μg/g

    Techniques Used: Mouse Assay

    IL-23 is elevated in vitro following hypoxia in a mouse hepatocyte and nonparenchymal cell (HC/NPC) coculture system. A : IL-23 mRNA levels as measured by p19 and p40 subunits in HC/NPC coculture system following hypoxia (1% O 2 ). B : Western blot showing
    Figure Legend Snippet: IL-23 is elevated in vitro following hypoxia in a mouse hepatocyte and nonparenchymal cell (HC/NPC) coculture system. A : IL-23 mRNA levels as measured by p19 and p40 subunits in HC/NPC coculture system following hypoxia (1% O 2 ). B : Western blot showing

    Techniques Used: In Vitro, Western Blot

    Overexpression of IL-23 in mice through adenovirus vector leads to induction of IL-17, interferon regulatory factor-1 (IRF-1), polymorphonuclear neutrophil (PMN) recruitment, and apoptosis. A : Western blot for IL-17 protein expression in the liver following
    Figure Legend Snippet: Overexpression of IL-23 in mice through adenovirus vector leads to induction of IL-17, interferon regulatory factor-1 (IRF-1), polymorphonuclear neutrophil (PMN) recruitment, and apoptosis. A : Western blot for IL-17 protein expression in the liver following

    Techniques Used: Over Expression, Mouse Assay, Plasmid Preparation, Western Blot, Expressing

    IL-23, interferon regulatory factor-1 (IRF-1), IL-17, and chemokine (C-X-C motif) ligand 2 (CXCL2) are elevated during cold ischemia-reperfusion (I/R) injury in mice. A : quantitative RT-PCR (qRT-PCR) was used to determine mRNA levels of IL-23 in liver
    Figure Legend Snippet: IL-23, interferon regulatory factor-1 (IRF-1), IL-17, and chemokine (C-X-C motif) ligand 2 (CXCL2) are elevated during cold ischemia-reperfusion (I/R) injury in mice. A : quantitative RT-PCR (qRT-PCR) was used to determine mRNA levels of IL-23 in liver

    Techniques Used: Mouse Assay, Quantitative RT-PCR

    Liver damage is attenuated during cold ischemia-reperfusion (I/R) injury by treatment with anti-IL-23 antibody. A and B : aspartate aminotransferase (AST)/alanine aminotransferase (ALT) serum levels at 6 h and 24 h following cold I/R with 18 h cold storage
    Figure Legend Snippet: Liver damage is attenuated during cold ischemia-reperfusion (I/R) injury by treatment with anti-IL-23 antibody. A and B : aspartate aminotransferase (AST)/alanine aminotransferase (ALT) serum levels at 6 h and 24 h following cold I/R with 18 h cold storage

    Techniques Used: AST Assay

    7) Product Images from "Innate Immune Responses to Systemic Acinetobacter baumannii Infection in Mice: Neutrophils, but Not Interleukin-17, Mediate Host Resistance ▿"

    Article Title: Innate Immune Responses to Systemic Acinetobacter baumannii Infection in Mice: Neutrophils, but Not Interleukin-17, Mediate Host Resistance ▿

    Journal: Infection and Immunity

    doi: 10.1128/IAI.00069-11

    In vitro induction of IL-17 from peritoneal exudate cells by recombinant IL-23 and LPS. PECs were collected from naive C3HeB/FeJ mice or from mice infected with a 1.2-LD 50 dose of A. baumannii strain 4502. Cells plated at the given concentrations were
    Figure Legend Snippet: In vitro induction of IL-17 from peritoneal exudate cells by recombinant IL-23 and LPS. PECs were collected from naive C3HeB/FeJ mice or from mice infected with a 1.2-LD 50 dose of A. baumannii strain 4502. Cells plated at the given concentrations were

    Techniques Used: In Vitro, Recombinant, Mouse Assay, Infection

    8) Product Images from "Excessive Polyamine Generation in Keratinocytes Promotes Self-RNA Endosomal Sensing by Dendritic Cells in Psoriasis"

    Article Title: Excessive Polyamine Generation in Keratinocytes Promotes Self-RNA Endosomal Sensing by Dendritic Cells in Psoriasis

    Journal: bioRxiv

    doi: 10.1101/2020.03.09.984658

    PP6 is diminished in the epidermis of psoriasis and its mouse models. ( A ) Immunohistochemical staining for PP6 in five healthy and five psoriatic human skin sections. Nor, normal skin; Pso, psoriatic skin. ( B ) Linear regression analysis of PP6 expression levels and acanthosis of 117 psoriatic skin biopsies (x axis: PP6 staining score; y axis: acanthosis of the skin section; r = -0.603). ( C ) The 117 psoriatic skin samples were separated into three groups according to the patient PASI. The PP6 staining score was compared between these groups. ( D and E ) Immunoblotting and immunohistochemical analysis of Pp6 expression in mouse skin derived from mice treated with vehicle (Ctr) or IMQ (n = 4). ( F and G ) Immunoblotting and immunohistochemical analysis of Pp6 expression in mouse ears injected with PBS (Ctr) or IL-23 (n = 2). For ( A , E and G) , the dotted lines indicate the border between the epidermis and the dermis; e, epidermis; d, dermis; scale bars represent 50 μm ( A ) or 20 μm ( E and G ). The data ( D - G ) are representative of three independent experiments. * p
    Figure Legend Snippet: PP6 is diminished in the epidermis of psoriasis and its mouse models. ( A ) Immunohistochemical staining for PP6 in five healthy and five psoriatic human skin sections. Nor, normal skin; Pso, psoriatic skin. ( B ) Linear regression analysis of PP6 expression levels and acanthosis of 117 psoriatic skin biopsies (x axis: PP6 staining score; y axis: acanthosis of the skin section; r = -0.603). ( C ) The 117 psoriatic skin samples were separated into three groups according to the patient PASI. The PP6 staining score was compared between these groups. ( D and E ) Immunoblotting and immunohistochemical analysis of Pp6 expression in mouse skin derived from mice treated with vehicle (Ctr) or IMQ (n = 4). ( F and G ) Immunoblotting and immunohistochemical analysis of Pp6 expression in mouse ears injected with PBS (Ctr) or IL-23 (n = 2). For ( A , E and G) , the dotted lines indicate the border between the epidermis and the dermis; e, epidermis; d, dermis; scale bars represent 50 μm ( A ) or 20 μm ( E and G ). The data ( D - G ) are representative of three independent experiments. * p

    Techniques Used: Immunohistochemistry, Staining, Expressing, Derivative Assay, Mouse Assay, Injection

    9) Product Images from "IL-36R signaling integrates innate and adaptive immune-mediated protection against enteropathogenic bacteria"

    Article Title: IL-36R signaling integrates innate and adaptive immune-mediated protection against enteropathogenic bacteria

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.2004484117

    IL-36R deficiency results in diminished IL-23, IL-6, and IL-22 expression in the large intestine during C. rodentium infection. ( A ) Experimental schematic of intestinal bacterial infection by oral infected Il1rl2 +/+ and Il1rl2 −/− mice with 5 to 6 × 10 9 CFU of C. rodentium . ( B and C ) PCR array gene expression analyses from large intestine tissues isolated at indicated time points (p.i.) of Il1rl2 +/+ and Il1rl2 −/− mice. ( D and E ) The top 10 significantly expressed genes with highest up-regulation were obtained from PCR array analysis between C. rodentium -infected Il1rl2 +/+ and Il1rl2 −/− mice at day 4 p.i. ( D ), and day 8 p.i. ( E ). ( F – H ) The time course of ( F ) IL-22, ( G ) IL-23, and ( H ) IL-6 protein expression from colonic tissues isolated from infected mice. Data are representative of two independent experiments with four to five mice per group. All data are presented as mean ± SEM (multiple t tests, one per row, corrected for multiple comparisons using the Holm–Sidak method; * P
    Figure Legend Snippet: IL-36R deficiency results in diminished IL-23, IL-6, and IL-22 expression in the large intestine during C. rodentium infection. ( A ) Experimental schematic of intestinal bacterial infection by oral infected Il1rl2 +/+ and Il1rl2 −/− mice with 5 to 6 × 10 9 CFU of C. rodentium . ( B and C ) PCR array gene expression analyses from large intestine tissues isolated at indicated time points (p.i.) of Il1rl2 +/+ and Il1rl2 −/− mice. ( D and E ) The top 10 significantly expressed genes with highest up-regulation were obtained from PCR array analysis between C. rodentium -infected Il1rl2 +/+ and Il1rl2 −/− mice at day 4 p.i. ( D ), and day 8 p.i. ( E ). ( F – H ) The time course of ( F ) IL-22, ( G ) IL-23, and ( H ) IL-6 protein expression from colonic tissues isolated from infected mice. Data are representative of two independent experiments with four to five mice per group. All data are presented as mean ± SEM (multiple t tests, one per row, corrected for multiple comparisons using the Holm–Sidak method; * P

    Techniques Used: Expressing, Infection, Mouse Assay, Polymerase Chain Reaction, Isolation

    Early IL-23 administration rescues protective immunity in C. rodentium -infected Il1rl2 −/− mice. ( A ) Experimental schematic of C. rodentium infection (5 to 6 × 10 9 CFU) by gastric gavage into Il1rl2 +/+ and Il1rl2 −/− mice, in the presence or absence of IL-23. ( B ) Serial whole-body imaging of infected mice at indicated time points. Images are representative of two independent experiments with at least five mice/group. ( C – E ) ( C ) Survival rate; ( D ) average body weight change; and ( E ) bacterial shedding in feces of infected mice at indicated time points. ( F and G ) ( F ) The H E staining and histology scoring of ( G ) colon sections from infected mice as in A are shown. ( H ) IL-22 protein expression in the colon at 10 days post C. rodentium infection from Il1rl2 +/+ and Il1rl2 −/− mice. ( I ) Colonic lamina propria cells of C. rodentium -infected mice at day 4 p.i. were isolated and analyzed by FACS for expression of intracellular IL-22 by Thy1 + RORγt + gated cells. ( J ) FACS frequency data of Thy1 + RORγt + IL-22 + gated cells of infected mice generated as in I . ( K – M ) ( K ) Claudin-2, ( L ) antimicrobial peptides, and ( M ) Mucin-2 mRNA expression in colon from infected mice at day 4 p.i. ( N ) Serial whole-body imaging of C. rodentium -infected Il1rl2 −/− mice in the presence or absence of IL-23 and neutralization antibodies, αCD90 or αCD4 as indicated. Images are representative of two independent experiments. ( O – P ) ( O ) Survival rate; ( P ) average body weight changes; and ( Q ) bacterial shedding in feces of infected mice as in N at indicated time points. ( R ) IL-22 protein determined by ELISA of colon of infected mice as in N at indicated time points. Data are representative of three independent experiments with five mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test. * P
    Figure Legend Snippet: Early IL-23 administration rescues protective immunity in C. rodentium -infected Il1rl2 −/− mice. ( A ) Experimental schematic of C. rodentium infection (5 to 6 × 10 9 CFU) by gastric gavage into Il1rl2 +/+ and Il1rl2 −/− mice, in the presence or absence of IL-23. ( B ) Serial whole-body imaging of infected mice at indicated time points. Images are representative of two independent experiments with at least five mice/group. ( C – E ) ( C ) Survival rate; ( D ) average body weight change; and ( E ) bacterial shedding in feces of infected mice at indicated time points. ( F and G ) ( F ) The H E staining and histology scoring of ( G ) colon sections from infected mice as in A are shown. ( H ) IL-22 protein expression in the colon at 10 days post C. rodentium infection from Il1rl2 +/+ and Il1rl2 −/− mice. ( I ) Colonic lamina propria cells of C. rodentium -infected mice at day 4 p.i. were isolated and analyzed by FACS for expression of intracellular IL-22 by Thy1 + RORγt + gated cells. ( J ) FACS frequency data of Thy1 + RORγt + IL-22 + gated cells of infected mice generated as in I . ( K – M ) ( K ) Claudin-2, ( L ) antimicrobial peptides, and ( M ) Mucin-2 mRNA expression in colon from infected mice at day 4 p.i. ( N ) Serial whole-body imaging of C. rodentium -infected Il1rl2 −/− mice in the presence or absence of IL-23 and neutralization antibodies, αCD90 or αCD4 as indicated. Images are representative of two independent experiments. ( O – P ) ( O ) Survival rate; ( P ) average body weight changes; and ( Q ) bacterial shedding in feces of infected mice as in N at indicated time points. ( R ) IL-22 protein determined by ELISA of colon of infected mice as in N at indicated time points. Data are representative of three independent experiments with five mice per group. All data are presented as mean ± SEM (one-way ANOVA with Tukey’s multiple comparison test. * P

    Techniques Used: Infection, Mouse Assay, Imaging, Staining, Expressing, Isolation, FACS, Generated, Neutralization, Enzyme-linked Immunosorbent Assay

    10) Product Images from "Resolvin E1 attenuates murine psoriatic dermatitis"

    Article Title: Resolvin E1 attenuates murine psoriatic dermatitis

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-30373-1

    RvE1 inhibits the production of IL-23 by DCs and migration of DCs and γδ T cells. ( a ) The effects of LTB4 (10 nM), RvE1(100 nM), and a BLT1 antagonist (U-75302) (10 nM) on IL-23p19 production by bone marrow-derived DCs. The amount of IL-23p19 in culture supernatant was measured by ELISA. ( b , c ) A representative FACS plot of Kaede red + cells ( b ) gated on DCs in Kaede transgenic mice, and the number ( c ) in dLNs of Kaede transgenic mice after IMQ application with or without RvE1 treatment. ( d , e ) Representative FACS plots of Kaede red + cells ( d ) gated on γδTCR mid+ cells and the number ( e ) in dLNs after IMQ application with or without RvE1 treatment. (f and g) Representative FACS plots of Kaede red + cells ( f ) gated on γδTCR mid+ cells, and the number ( g ) in dLNs of Kaede transgenic mice treated with or without RvE1 treatment in the steady state. ( h , i ) Transwell migration assay for γδTCR mid+ cells ( h ) and BMDCs ( i ). Thirty minutes after RvE1 (100 nM), a BLT1 antagonist (10 nM) or vehicle treatment, cells were tested for transmigration for 3 h (γδTCR mid+ cells) or 5 h (BMDCs) in the presence of vehicle or LTB4 (10 nM) in the lower chamber. Results are expressed as the mean ± SD. All p -values were obtained by Student’s t- test: * p
    Figure Legend Snippet: RvE1 inhibits the production of IL-23 by DCs and migration of DCs and γδ T cells. ( a ) The effects of LTB4 (10 nM), RvE1(100 nM), and a BLT1 antagonist (U-75302) (10 nM) on IL-23p19 production by bone marrow-derived DCs. The amount of IL-23p19 in culture supernatant was measured by ELISA. ( b , c ) A representative FACS plot of Kaede red + cells ( b ) gated on DCs in Kaede transgenic mice, and the number ( c ) in dLNs of Kaede transgenic mice after IMQ application with or without RvE1 treatment. ( d , e ) Representative FACS plots of Kaede red + cells ( d ) gated on γδTCR mid+ cells and the number ( e ) in dLNs after IMQ application with or without RvE1 treatment. (f and g) Representative FACS plots of Kaede red + cells ( f ) gated on γδTCR mid+ cells, and the number ( g ) in dLNs of Kaede transgenic mice treated with or without RvE1 treatment in the steady state. ( h , i ) Transwell migration assay for γδTCR mid+ cells ( h ) and BMDCs ( i ). Thirty minutes after RvE1 (100 nM), a BLT1 antagonist (10 nM) or vehicle treatment, cells were tested for transmigration for 3 h (γδTCR mid+ cells) or 5 h (BMDCs) in the presence of vehicle or LTB4 (10 nM) in the lower chamber. Results are expressed as the mean ± SD. All p -values were obtained by Student’s t- test: * p

    Techniques Used: Migration, Derivative Assay, Enzyme-linked Immunosorbent Assay, FACS, Transgenic Assay, Mouse Assay, Transwell Migration Assay, Transmigration Assay

    Gene expressions in human psoriatic skin, and inhibitory effects of RvE1 on the IL-23 production by DCs and migration of DCs, Th17 and Tc17 cells. ( a ) Box and whisker plots of mRNA levels of IL23A , IL17A , LTB4R , LTA4H , and ALOX5 . Box and whisker plots of Z scores ( Y -axis) for skin taken from human healthy controls, and non-lesional and lesional skin taken from psoriasis patients in a public data set (GEO accession no. GSE13355). The length of the box represents the distance between 25% and 75%, and the horizontal line inside the box represents the group median. The whiskers indicate minimum and maximum values. All p -values were obtained by nonparameteric Wilcoxon-Mann-Whitney test. ( b ) The effects of LTB4 (100 nM), RvE1 (100 nM) and a BLT1 antagonist (10 nM) on IL-23p19 production by human monocyte-derived DCs. The amount of IL-23p19 in culture supernatant was measured by ELISA. ( c ) Transwell migration assay for human MoDCs, Tc17, and Th17 cells. Thirty minutes after RvE1 (100 nM), a BLT1 antagonist (U-75302) (10 nM), or vehicle treatment, human MoDCs, Tc17 and Th17 cells were tested for transmigration for 5 h (BMDCs) or 3 h (Tc17 and Th17 cells) in the presence of vehicle or LTB4 (10 nM) in the lower (BMDCs) or upper (Tc17 and Th17 cells) chamber. Results are expressed as the mean ± SD. All p -values were obtained by Student’s t- test: * p
    Figure Legend Snippet: Gene expressions in human psoriatic skin, and inhibitory effects of RvE1 on the IL-23 production by DCs and migration of DCs, Th17 and Tc17 cells. ( a ) Box and whisker plots of mRNA levels of IL23A , IL17A , LTB4R , LTA4H , and ALOX5 . Box and whisker plots of Z scores ( Y -axis) for skin taken from human healthy controls, and non-lesional and lesional skin taken from psoriasis patients in a public data set (GEO accession no. GSE13355). The length of the box represents the distance between 25% and 75%, and the horizontal line inside the box represents the group median. The whiskers indicate minimum and maximum values. All p -values were obtained by nonparameteric Wilcoxon-Mann-Whitney test. ( b ) The effects of LTB4 (100 nM), RvE1 (100 nM) and a BLT1 antagonist (10 nM) on IL-23p19 production by human monocyte-derived DCs. The amount of IL-23p19 in culture supernatant was measured by ELISA. ( c ) Transwell migration assay for human MoDCs, Tc17, and Th17 cells. Thirty minutes after RvE1 (100 nM), a BLT1 antagonist (U-75302) (10 nM), or vehicle treatment, human MoDCs, Tc17 and Th17 cells were tested for transmigration for 5 h (BMDCs) or 3 h (Tc17 and Th17 cells) in the presence of vehicle or LTB4 (10 nM) in the lower (BMDCs) or upper (Tc17 and Th17 cells) chamber. Results are expressed as the mean ± SD. All p -values were obtained by Student’s t- test: * p

    Techniques Used: Migration, Whisker Assay, MANN-WHITNEY, Derivative Assay, Enzyme-linked Immunosorbent Assay, Transwell Migration Assay, Transmigration Assay

    11) Product Images from "Resolvin E1 attenuates murine psoriatic dermatitis"

    Article Title: Resolvin E1 attenuates murine psoriatic dermatitis

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-30373-1

    RvE1 inhibits the production of IL-23 by DCs and migration of DCs and γδ T cells. ( a ) The effects of LTB4 (10 nM), RvE1(100 nM), and a BLT1 antagonist (U-75302) (10 nM) on IL-23p19 production by bone marrow-derived DCs. The amount of IL-23p19 in culture supernatant was measured by ELISA. ( b , c ) A representative FACS plot of Kaede red + cells ( b ) gated on DCs in Kaede transgenic mice, and the number ( c ) in dLNs of Kaede transgenic mice after IMQ application with or without RvE1 treatment. ( d , e ) Representative FACS plots of Kaede red + cells ( d ) gated on γδTCR mid+ cells and the number ( e ) in dLNs after IMQ application with or without RvE1 treatment. (f and g) Representative FACS plots of Kaede red + cells ( f ) gated on γδTCR mid+ cells, and the number ( g ) in dLNs of Kaede transgenic mice treated with or without RvE1 treatment in the steady state. ( h , i ) Transwell migration assay for γδTCR mid+ cells ( h ) and BMDCs ( i ). Thirty minutes after RvE1 (100 nM), a BLT1 antagonist (10 nM) or vehicle treatment, cells were tested for transmigration for 3 h (γδTCR mid+ cells) or 5 h (BMDCs) in the presence of vehicle or LTB4 (10 nM) in the lower chamber. Results are expressed as the mean ± SD. All p -values were obtained by Student’s t- test: * p
    Figure Legend Snippet: RvE1 inhibits the production of IL-23 by DCs and migration of DCs and γδ T cells. ( a ) The effects of LTB4 (10 nM), RvE1(100 nM), and a BLT1 antagonist (U-75302) (10 nM) on IL-23p19 production by bone marrow-derived DCs. The amount of IL-23p19 in culture supernatant was measured by ELISA. ( b , c ) A representative FACS plot of Kaede red + cells ( b ) gated on DCs in Kaede transgenic mice, and the number ( c ) in dLNs of Kaede transgenic mice after IMQ application with or without RvE1 treatment. ( d , e ) Representative FACS plots of Kaede red + cells ( d ) gated on γδTCR mid+ cells and the number ( e ) in dLNs after IMQ application with or without RvE1 treatment. (f and g) Representative FACS plots of Kaede red + cells ( f ) gated on γδTCR mid+ cells, and the number ( g ) in dLNs of Kaede transgenic mice treated with or without RvE1 treatment in the steady state. ( h , i ) Transwell migration assay for γδTCR mid+ cells ( h ) and BMDCs ( i ). Thirty minutes after RvE1 (100 nM), a BLT1 antagonist (10 nM) or vehicle treatment, cells were tested for transmigration for 3 h (γδTCR mid+ cells) or 5 h (BMDCs) in the presence of vehicle or LTB4 (10 nM) in the lower chamber. Results are expressed as the mean ± SD. All p -values were obtained by Student’s t- test: * p

    Techniques Used: Migration, Derivative Assay, Enzyme-linked Immunosorbent Assay, FACS, Transgenic Assay, Mouse Assay, Transwell Migration Assay, Transmigration Assay

    12) Product Images from "IL-22 is required for the induction of bronchus-associated lymphoid tissue in tolerant lung allografts"

    Article Title: IL-22 is required for the induction of bronchus-associated lymphoid tissue in tolerant lung allografts

    Journal: American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons

    doi: 10.1111/ajt.15701

    Late reconstitution of IL-22 expression mediates B cell recruitment to lymphoid follicles. Histological appearance of Balb/c lung grafts initially transplanted into IL-22 deficient recipients that received peri-operative costimulatory blockade and then retransplanted into non-immunosuppressed wildtype secondary recipients 30 days later. Images show lung allografts harvested (A) 7 and (B) 30 days after retransplantation. n=3. Scale bars =100 μm. Quantification of lymphoid follicle (C) number and (D) percent of total area per section in IL-22-deficient recipients 30 days after primary transplantation, as well as 7 and 30 days after retransplantation into wildtype secondary hosts. Histological appearance of B cell (green), T cell (red), and DAPI (blue) immunofluorescent staining from Balb/c lung grafts (E) 7 and (F) 30 days after retransplantation (scale bars =50 μm) and (G) quantification of B220 / CD3 cell ratio per lymphoid follicle among the various conditions. Each dot represents one lymphoid follicle (n≥2 mice per group). Immunostaining for Foxp3 (brown) (H) 30 days after retransplantation and (I) quantification of Foxp3 cells per lymphoid follicle at 30 days among various conditions. Scale bars =50 μm. Each dot represents one lymphoid follicle (n≥2 mice per group). Immunostaining for PNAd (brown) (J) 30 days after retransplantation. Scale bars =50 μm. (K) Histological appearance of Balb/c lung grafts initially transplanted into IL-23 neutralized B6 Foxp3-DTR recipients that received peri-operative costimulatory blockade and then retransplanted into non-immunosuppressed wildtype B6 recipients 30 days later that received DT at the time of retransplantation. Images are shown 7 days after retransplantation (H E) (n=4). Scale bars =100 μm. ns= not significant; *p
    Figure Legend Snippet: Late reconstitution of IL-22 expression mediates B cell recruitment to lymphoid follicles. Histological appearance of Balb/c lung grafts initially transplanted into IL-22 deficient recipients that received peri-operative costimulatory blockade and then retransplanted into non-immunosuppressed wildtype secondary recipients 30 days later. Images show lung allografts harvested (A) 7 and (B) 30 days after retransplantation. n=3. Scale bars =100 μm. Quantification of lymphoid follicle (C) number and (D) percent of total area per section in IL-22-deficient recipients 30 days after primary transplantation, as well as 7 and 30 days after retransplantation into wildtype secondary hosts. Histological appearance of B cell (green), T cell (red), and DAPI (blue) immunofluorescent staining from Balb/c lung grafts (E) 7 and (F) 30 days after retransplantation (scale bars =50 μm) and (G) quantification of B220 / CD3 cell ratio per lymphoid follicle among the various conditions. Each dot represents one lymphoid follicle (n≥2 mice per group). Immunostaining for Foxp3 (brown) (H) 30 days after retransplantation and (I) quantification of Foxp3 cells per lymphoid follicle at 30 days among various conditions. Scale bars =50 μm. Each dot represents one lymphoid follicle (n≥2 mice per group). Immunostaining for PNAd (brown) (J) 30 days after retransplantation. Scale bars =50 μm. (K) Histological appearance of Balb/c lung grafts initially transplanted into IL-23 neutralized B6 Foxp3-DTR recipients that received peri-operative costimulatory blockade and then retransplanted into non-immunosuppressed wildtype B6 recipients 30 days later that received DT at the time of retransplantation. Images are shown 7 days after retransplantation (H E) (n=4). Scale bars =100 μm. ns= not significant; *p

    Techniques Used: Expressing, Transplantation Assay, Staining, Mouse Assay, Immunostaining

    BALT induction in tolerant lung allografts requires IL-22 and IL-23 expression. Histological appearance of Balb/c lung allografts (A) 7 and (B) 30 days after transplantation into B6 wildtype (WT) mice that received peri-operative costimulatory blockade, with arrow depicting BALT at 30 days. Relative RNA expression of (C) IL-22, (D) IL-23, (E) LTβ, and (F) IL-17 from whole lung lysates of naïve Balb/c lungs and transplanted lungs at days 7 and 30. Appearance of lung allografts 30 days after transplantation into (G) IL-22 deficient hosts or (H) wildtype recipients receiving IL-23 neutralization. Recipients were treated with peri-operative costimulatory blockade. Scale bars: 100μM. (I) Quantification of the number of lymphoid follicles per section 30 days after transplantation into various recipients. n≥4 for all conditions. * p
    Figure Legend Snippet: BALT induction in tolerant lung allografts requires IL-22 and IL-23 expression. Histological appearance of Balb/c lung allografts (A) 7 and (B) 30 days after transplantation into B6 wildtype (WT) mice that received peri-operative costimulatory blockade, with arrow depicting BALT at 30 days. Relative RNA expression of (C) IL-22, (D) IL-23, (E) LTβ, and (F) IL-17 from whole lung lysates of naïve Balb/c lungs and transplanted lungs at days 7 and 30. Appearance of lung allografts 30 days after transplantation into (G) IL-22 deficient hosts or (H) wildtype recipients receiving IL-23 neutralization. Recipients were treated with peri-operative costimulatory blockade. Scale bars: 100μM. (I) Quantification of the number of lymphoid follicles per section 30 days after transplantation into various recipients. n≥4 for all conditions. * p

    Techniques Used: Expressing, Transplantation Assay, Mouse Assay, RNA Expression, Neutralization

    IL-22 is required for B cell, but not Foxp3 + cell recruitment into lymphoid follicles. Immunofluorescent staining of B cells (green), T cells (red), and DAPI (blue) in Balb/c lung allografts 30 days after transplantation into (A) wildtype (WT), (B) IL-22-deficient, (C) IL-23-neutralized, (D) AR mice (RORγt-cre Ahr fl/fl ) and (E) γδ T cell-deficient (TCRδ −/− ) recipients. Scale bars: 100 μm (F) Quantification of B220 / CD3 positive cell ratio per lymphoid follicle 30 days after transplantation into the indicated recipients. Each dot represents one lymphoid follicle (n≥2 mice per group). Immunostaining for Foxp3 (brown) in Balb/c lung allografts 30 days after transplantation into (G) wildtype, (H) IL-22 deficient, (I) IL-23 neutralized, (J) AR mice, and (K) γδ T cell-deficient recipients. (L) Quantification of Foxp3 cells per lymphoid follicle 30 days after transplantation of Balb/c lungs into the indicated recipients. Each dot represents one lymphoid follicle (n≥2 mice per group). Recipients were treated with peri-operative co-stimulatory blockade for all conditions. Scale bars: 50 μm.
    Figure Legend Snippet: IL-22 is required for B cell, but not Foxp3 + cell recruitment into lymphoid follicles. Immunofluorescent staining of B cells (green), T cells (red), and DAPI (blue) in Balb/c lung allografts 30 days after transplantation into (A) wildtype (WT), (B) IL-22-deficient, (C) IL-23-neutralized, (D) AR mice (RORγt-cre Ahr fl/fl ) and (E) γδ T cell-deficient (TCRδ −/− ) recipients. Scale bars: 100 μm (F) Quantification of B220 / CD3 positive cell ratio per lymphoid follicle 30 days after transplantation into the indicated recipients. Each dot represents one lymphoid follicle (n≥2 mice per group). Immunostaining for Foxp3 (brown) in Balb/c lung allografts 30 days after transplantation into (G) wildtype, (H) IL-22 deficient, (I) IL-23 neutralized, (J) AR mice, and (K) γδ T cell-deficient recipients. (L) Quantification of Foxp3 cells per lymphoid follicle 30 days after transplantation of Balb/c lungs into the indicated recipients. Each dot represents one lymphoid follicle (n≥2 mice per group). Recipients were treated with peri-operative co-stimulatory blockade for all conditions. Scale bars: 50 μm.

    Techniques Used: Staining, Transplantation Assay, Mouse Assay, Immunostaining

    Expression of peripheral node addressin in lymphoid follicles is dependent on IL-22. Histological appearance and representative staining for PNAd (brown) in Balb/c lung allografts 30 days after transplantation into (A) wildtype (WT), (B) IL-22-deficient, (C) IL-23-neutralized, (D) AR (RORγt-cre Ahr fl/fl ) mice, and (E) γδ T cell-deficient (TCRδ −/− ) recipients. Recipient mice were treated with peri-operative costimulatory blockade. Scale bars: 50 μm.
    Figure Legend Snippet: Expression of peripheral node addressin in lymphoid follicles is dependent on IL-22. Histological appearance and representative staining for PNAd (brown) in Balb/c lung allografts 30 days after transplantation into (A) wildtype (WT), (B) IL-22-deficient, (C) IL-23-neutralized, (D) AR (RORγt-cre Ahr fl/fl ) mice, and (E) γδ T cell-deficient (TCRδ −/− ) recipients. Recipient mice were treated with peri-operative costimulatory blockade. Scale bars: 50 μm.

    Techniques Used: Expressing, Staining, Transplantation Assay, Mouse Assay

    13) Product Images from "CCR6 is required for epidermal trafficking of ?? T cells in an IL-23-induced model of psoriasiform dermatitis"

    Article Title: CCR6 is required for epidermal trafficking of ?? T cells in an IL-23-induced model of psoriasiform dermatitis

    Journal: The Journal of investigative dermatology

    doi: 10.1038/jid.2012.260

    Neutralizing anti-CCL20 monoclonal antibodies inhibit the trafficking of GDL T cells in response to IL-23 injection Following treatment with neutralizing anti-CCL20 mAb, isotype control, or PBS injection for 5 days, both pooled epidermal ( a ) and dermal ( b ) cell suspensions from 4 mouse ears (per condition) were stained with mAbs against γδ TCR for flow cytometry e( a,b ) or processed for RNA extraction for RT-PCR ( c,d ). In ( a,b ), the percentage indicated in each dot plot reflects the number of GDL cells (i.e., events) in the oval window divided by the total sample events shown in the entire dot plot. Percent reduction in epidermal GDL T cells was calculated at 76% for data shown [(% GDL cells IL23 + isotype)-(% GDL cells IL23+anti-CCL20)]/[(%GDL cells IL23+isotype)-(%GDL cells PBS)]*100. Pooled epidermal ( c ) and dermal ( d ) cell suspensions (see above) were used for quantitative RT-PCR to measure expression of IL-22 and CCR6. Fold changes were calculated for IL-22 and Ccr6 mRNAs normalized for Gapdh mRNA vs . the PBS treated sample. Error bars indicate SD ( c,d ) based on a single, pooled treatment group that was divided into three wells for RT-PCR measurement. Similar results were obtained in 2 independent experiments.
    Figure Legend Snippet: Neutralizing anti-CCL20 monoclonal antibodies inhibit the trafficking of GDL T cells in response to IL-23 injection Following treatment with neutralizing anti-CCL20 mAb, isotype control, or PBS injection for 5 days, both pooled epidermal ( a ) and dermal ( b ) cell suspensions from 4 mouse ears (per condition) were stained with mAbs against γδ TCR for flow cytometry e( a,b ) or processed for RNA extraction for RT-PCR ( c,d ). In ( a,b ), the percentage indicated in each dot plot reflects the number of GDL cells (i.e., events) in the oval window divided by the total sample events shown in the entire dot plot. Percent reduction in epidermal GDL T cells was calculated at 76% for data shown [(% GDL cells IL23 + isotype)-(% GDL cells IL23+anti-CCL20)]/[(%GDL cells IL23+isotype)-(%GDL cells PBS)]*100. Pooled epidermal ( c ) and dermal ( d ) cell suspensions (see above) were used for quantitative RT-PCR to measure expression of IL-22 and CCR6. Fold changes were calculated for IL-22 and Ccr6 mRNAs normalized for Gapdh mRNA vs . the PBS treated sample. Error bars indicate SD ( c,d ) based on a single, pooled treatment group that was divided into three wells for RT-PCR measurement. Similar results were obtained in 2 independent experiments.

    Techniques Used: Injection, Staining, Flow Cytometry, Cytometry, RNA Extraction, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Expressing

    Neutralizing anti-CCL20 monoclonal antibodies inhibit the ear swelling in response to IL-23 injection ( a ) WT mice were injected with neutralizing anti-CCL20 mAb, isotype control, or PBS in ear skin with subsequent measurement of ear thickness throughout the treatment period. *P
    Figure Legend Snippet: Neutralizing anti-CCL20 monoclonal antibodies inhibit the ear swelling in response to IL-23 injection ( a ) WT mice were injected with neutralizing anti-CCL20 mAb, isotype control, or PBS in ear skin with subsequent measurement of ear thickness throughout the treatment period. *P

    Techniques Used: Injection, Mouse Assay

    CCL20 expression in mouse epidermis is upregulated within 24 hr of IL-23 injection IL-23 was injected into ears of WT mice at day 0, 2, and 4. Mice were euthanized on the indicated days and ears were stained with anti-CCL20 antibody (red) and DAPI (nuclear counterstain in blue). Isotype control staining showed no epidermal staining with either IL-23 or PBS-injected ears (data not shown). Scale bar = 50 μm.
    Figure Legend Snippet: CCL20 expression in mouse epidermis is upregulated within 24 hr of IL-23 injection IL-23 was injected into ears of WT mice at day 0, 2, and 4. Mice were euthanized on the indicated days and ears were stained with anti-CCL20 antibody (red) and DAPI (nuclear counterstain in blue). Isotype control staining showed no epidermal staining with either IL-23 or PBS-injected ears (data not shown). Scale bar = 50 μm.

    Techniques Used: Expressing, Injection, Mouse Assay, Staining

    Immunofluorescence staining for CCR6 expression following IL-23 injection Following IL-23 ( a ) or PBS ( b ) injection for 6 days, mice were euthanized and ears were stained with anti-CCR6 mAbs (green) and anti-Laminin 332 mAbs (basement membrane in red), and the 3D images were acquired using a confocal microscope. The figure shows the perspective from the dermis toward the epidermis. The width of basement membrane in red shows the depth of the sections. Larger numbers of CCR6-expressing cells in the epidermis (above the basement membrane) were revealed in IL-23-, but not PBS-injected, mice. CCR6-expressing cells ( a ) at (arrow head), immediately adjacent to (double arrowheads), or above (arrow) the basement membrane in the dashed rectangular area marked in the upper figure are shown at higher magnification below. Isotype control staining showed no staining with either IL-23 or PBS-injected ears (data not shown). Scale bar = 10 μm. In ( c ), IL-23-treated skin sections were stained with labeled mAbs against CCR6 (green) and the γδ-TCR (red). Images were acquired by confocal microscopy at the level of the epidermis (x63 objective, scale bar as indicated). Similar results were obtained in 3 independent experiments.
    Figure Legend Snippet: Immunofluorescence staining for CCR6 expression following IL-23 injection Following IL-23 ( a ) or PBS ( b ) injection for 6 days, mice were euthanized and ears were stained with anti-CCR6 mAbs (green) and anti-Laminin 332 mAbs (basement membrane in red), and the 3D images were acquired using a confocal microscope. The figure shows the perspective from the dermis toward the epidermis. The width of basement membrane in red shows the depth of the sections. Larger numbers of CCR6-expressing cells in the epidermis (above the basement membrane) were revealed in IL-23-, but not PBS-injected, mice. CCR6-expressing cells ( a ) at (arrow head), immediately adjacent to (double arrowheads), or above (arrow) the basement membrane in the dashed rectangular area marked in the upper figure are shown at higher magnification below. Isotype control staining showed no staining with either IL-23 or PBS-injected ears (data not shown). Scale bar = 10 μm. In ( c ), IL-23-treated skin sections were stained with labeled mAbs against CCR6 (green) and the γδ-TCR (red). Images were acquired by confocal microscopy at the level of the epidermis (x63 objective, scale bar as indicated). Similar results were obtained in 3 independent experiments.

    Techniques Used: Immunofluorescence, Staining, Expressing, Injection, Mouse Assay, Microscopy, Labeling, Confocal Microscopy

    CCR6 KO mice fail to accumulate IL-22 + , GDL T cells in response to IL-23 injection Following IL-23 injection for 6 days, both WT and CCR6 KO mice were euthanized. Both epidermal and dermal cell suspensions were analyzed by flow cytometry with γδ TCR, CD3, and IL-22 mAbs. ( a ) Total numbers of GDL T cells per PBS- or IL-23-injected ear of WT and CCR6 KO mice were counted. The percentages of CD3 + GDL T cells ( b ) and IL-22 + GDL T cells ( c ) were calculated. *P
    Figure Legend Snippet: CCR6 KO mice fail to accumulate IL-22 + , GDL T cells in response to IL-23 injection Following IL-23 injection for 6 days, both WT and CCR6 KO mice were euthanized. Both epidermal and dermal cell suspensions were analyzed by flow cytometry with γδ TCR, CD3, and IL-22 mAbs. ( a ) Total numbers of GDL T cells per PBS- or IL-23-injected ear of WT and CCR6 KO mice were counted. The percentages of CD3 + GDL T cells ( b ) and IL-22 + GDL T cells ( c ) were calculated. *P

    Techniques Used: Mouse Assay, Injection, Flow Cytometry, Cytometry

    Related Articles

    Recombinant:

    Article Title: IL-23 induced in keratinocytes by endogenous TLR4 ligands polarizes dendritic cells to drive IL-22 responses to skin immunization
    Article Snippet: Isolation and functional analysis of mouse skin and skin DLN DCs CD11c+ DCs of mouse skin and skin DLN (axillary and inguinal) were purified as described previously ( ). .. DCs were primed with SCM or recombinant IL-23 (20 ng/ml; R & D Systems) for 24 h. After extensive washing, DCs were co-cultured with naive CD4+ CD62L+ T cells isolated by using naive CD4 T cell isolation kit II (Miltenyi Biotec) in the presence of OVA323–339 peptide (4 µM; AnaSpec Inc.) for 4–5 d. In some experiments, neutralizing antibody goat anti–mouse IL-23 ( ) was used at 10 µg/ml. .. For in vivo DC priming experiment, OVA (1 mg in 100 µl saline) or saline was epicutaneously applied to shaved, tape-stripped skin of WT, Tlr4−/− , and Il23r−/− mice.

    Article Title: Lack of evidence for expression and function of IL-39 in human immune cells
    Article Snippet: After washing of the plates, biotinylated EBI3 antibody (Biorbyt, 3 μg/ml) was added for 1 hr, followed by washing and by incubation with HRP-conjugated Avidin (Biolegend). .. In order to determine whether the IL-39 ELISA recognizes other p19- or EBI3-containing cytokines, 200 nM recombinant IL-23 or IL-27 cytokines (R & D Systems) were added to the anti-p19 coated and blocked plates. .. After 2 hrs of incubation at room temperature, the plates were washed and biotinylated EBI3 antibody was added for 1 hr.

    Article Title: Prognostic value of serum soluble interleukin‐23 receptor and related T‐helper 17 cell cytokines in non‐small cell lung carcinoma, et al. Prognostic value of serum soluble interleukin‐23 receptor and related T‐helper 17 cell cytokines in non‐small cell lung carcinoma
    Article Snippet: The commercial ELISA kits of IL‐23 and IL‐17 (R & D Systems) was carried out according to the manufacturer’s instructions. .. 2.11 Serum soluble IL‐23R bound with IL‐23 in serum of NSCLC by ELISA assay Interleukin‐23 recombinant protein (100 μL) (800 pg 1887‐ML‐010; R & D Systems) was coated in the ELISA plate at 4°C overnight, with assay buffer (1× PBS and 3% BSA) as negative control. .. After washing with washing buffer (0.05% Tween‐20) 3 times, 100 μL serum samples with high (50‐100 pg/mL) or low (20‐50 pg/mL) levels of sIL‐23R validated by ELISA from 20 NSCLC patients were incubated for 2 hours at room temperature.

    Article Title: Resolvin E1 regulates interleukin-23, interferon-? and lipoxin A4 to promote resolution of allergic airway inflammation
    Article Snippet: To investigate inflammation resolution, some mice received RvE1 methyl ester (50 ng or 100 ng), a 15 epi-LXA4 analog (15-epi - 16 para-fluro-phenoxy-LXA4 methyl ester, ATLa) (100ng) or vehicle by intravenous injection on only days 18, 19 and 20. .. Some animals were treated with recombinant murine IL-17A (1μg/day) (eBiosciences), a neutralizing IL-17A-specific antibody (10 μg/day) (eBioMM17F3) or an isotype control antibody (eBiosciences), recombinant murine IL-23 (1μg/day) (R & D Systems), anti-IFN-γ (10 μg/day) (XMG1.2, Biolegend), or recombinant murine IL-6 (1μg/day) (PeproTech) by intranasal administration. ..

    Article Title: Cutting Edge: NKT Cells Constitutively Express IL-23 Receptor and RORγt and Rapidly Produce IL-17 upon Receptor Ligation in an IL-6-Independent Fashion 1
    Article Snippet: Cells were stimulated either by adding 2.5 μ g/ml anti-CD3 (clone 145−2C11) directly into cell culture or by using plates precoated with anti-CD3 at 10 μ g/ml. .. Where indicated, 10 ng/ml recombinant murine IL-23 was added (R & D Systems). α -GalCer was used at 100 ng/ml, presented by CD11c-positive cells isolated by autoMACS (Miltenyi Biotech). .. Mouse IL-17A and IFN- γ protein were detected in supernatants by DuoSet ELISA-matched Ab pairs (R & D Systems) as per the manufacturer's instructions.

    Article Title: Immune requirements for protective Th17 recall responses to Mycobacterium tuberculosis challenge
    Article Snippet: Adoptive T cell transfer and experimental infections Naïve T cells were isolated from ESAT-6 Tg mice using CD4+ (L3T4) magnetic bead sorting (Miltenyi Biotec, San Diego, CA). .. To generate Th17 cells, CD4+ T cells were cultured at a 1:1 ratio with BMDCs in the presence of ESAT-61-20 peptide (10 μg/mL), recombinant (r)-mouse IL-2 (10 U/mL), r-mouse IL-6 (30 ng/ml R & D Systems), r-mouse IL-23 (50 ng/ml R & D Systems), r-human TGF-β (5 ng/mL R & D Systems), anti-IL-4 antibody (10 μg/mL), and anti-IFN-γ antibody (10 μg/mL) in Iscove’s Modified Dulbecco’s Medium (Life Technologies, Grand Island, NY). .. For Th1 cells, CD4+ T cells were cultured at a 1:1 ratio with BMDCs in the presence of ESAT-61-20 peptide (10 μg/mL), r-mouse IL-2 (10 U/mL), r-mouse IL-12 (10 ng/ml R & D Systems) and anti-IL-4 antibody (10 μg/mL) in Iscove’s Modified Dulbecco’s Medium (Life Technologies, Grand Island, NY).

    Article Title: The Involvement of IL-17A in the Murine Response to Sub-Lethal Inhalational Infection with Francisella tularensis
    Article Snippet: The anti IL-17A antibodies (for flow cytometry and for in vivo neutralization) are specific to this isoform and do not cross react with other isoforms. .. Recombinant murine IL-17A was purchased from Prospec (Rehovot, Israel) and recombinant murine IL-23 was purchased from R & D Systems. ..

    Article Title: Microbiota-derived butyrate suppresses group 3 innate lymphoid cells in terminal ileal Peyer’s patches
    Article Snippet: For intracellular cytokine staining of Foxp3, cells were fixed, permeabilized, and stained with FITC-conjugated anti-CD4 Ab, PE-conjugated anti-CD25 Ab, and APC-conjugated anti-Foxp3 Ab using a Foxp3 staining set (eBioscience, San Diego, CA), according to the manufacturer’s protocol. .. For intracellular cytokine staining of IL-22, PP cells were cultured with or without butyrate and then stimulated for 4 hr with PMA (50 ng/mL), ionomycin (500 ng/mL), and mouse recombinant IL-23 (40 ng/mL; R & D Systems, Minneapolis, MN). .. Brefeldin A (eBioscience) was added to the culture medium and cells were incubated for the final 1.5 hr.

    Isolation:

    Article Title: IL-23 induced in keratinocytes by endogenous TLR4 ligands polarizes dendritic cells to drive IL-22 responses to skin immunization
    Article Snippet: Isolation and functional analysis of mouse skin and skin DLN DCs CD11c+ DCs of mouse skin and skin DLN (axillary and inguinal) were purified as described previously ( ). .. DCs were primed with SCM or recombinant IL-23 (20 ng/ml; R & D Systems) for 24 h. After extensive washing, DCs were co-cultured with naive CD4+ CD62L+ T cells isolated by using naive CD4 T cell isolation kit II (Miltenyi Biotec) in the presence of OVA323–339 peptide (4 µM; AnaSpec Inc.) for 4–5 d. In some experiments, neutralizing antibody goat anti–mouse IL-23 ( ) was used at 10 µg/ml. .. For in vivo DC priming experiment, OVA (1 mg in 100 µl saline) or saline was epicutaneously applied to shaved, tape-stripped skin of WT, Tlr4−/− , and Il23r−/− mice.

    Article Title: Cutting Edge: NKT Cells Constitutively Express IL-23 Receptor and RORγt and Rapidly Produce IL-17 upon Receptor Ligation in an IL-6-Independent Fashion 1
    Article Snippet: Cells were stimulated either by adding 2.5 μ g/ml anti-CD3 (clone 145−2C11) directly into cell culture or by using plates precoated with anti-CD3 at 10 μ g/ml. .. Where indicated, 10 ng/ml recombinant murine IL-23 was added (R & D Systems). α -GalCer was used at 100 ng/ml, presented by CD11c-positive cells isolated by autoMACS (Miltenyi Biotech). .. Mouse IL-17A and IFN- γ protein were detected in supernatants by DuoSet ELISA-matched Ab pairs (R & D Systems) as per the manufacturer's instructions.

    Cell Isolation:

    Article Title: IL-23 induced in keratinocytes by endogenous TLR4 ligands polarizes dendritic cells to drive IL-22 responses to skin immunization
    Article Snippet: Isolation and functional analysis of mouse skin and skin DLN DCs CD11c+ DCs of mouse skin and skin DLN (axillary and inguinal) were purified as described previously ( ). .. DCs were primed with SCM or recombinant IL-23 (20 ng/ml; R & D Systems) for 24 h. After extensive washing, DCs were co-cultured with naive CD4+ CD62L+ T cells isolated by using naive CD4 T cell isolation kit II (Miltenyi Biotec) in the presence of OVA323–339 peptide (4 µM; AnaSpec Inc.) for 4–5 d. In some experiments, neutralizing antibody goat anti–mouse IL-23 ( ) was used at 10 µg/ml. .. For in vivo DC priming experiment, OVA (1 mg in 100 µl saline) or saline was epicutaneously applied to shaved, tape-stripped skin of WT, Tlr4−/− , and Il23r−/− mice.

    Enzyme-linked Immunosorbent Assay:

    Article Title: Lack of evidence for expression and function of IL-39 in human immune cells
    Article Snippet: After washing of the plates, biotinylated EBI3 antibody (Biorbyt, 3 μg/ml) was added for 1 hr, followed by washing and by incubation with HRP-conjugated Avidin (Biolegend). .. In order to determine whether the IL-39 ELISA recognizes other p19- or EBI3-containing cytokines, 200 nM recombinant IL-23 or IL-27 cytokines (R & D Systems) were added to the anti-p19 coated and blocked plates. .. After 2 hrs of incubation at room temperature, the plates were washed and biotinylated EBI3 antibody was added for 1 hr.

    Article Title: Prognostic value of serum soluble interleukin‐23 receptor and related T‐helper 17 cell cytokines in non‐small cell lung carcinoma, et al. Prognostic value of serum soluble interleukin‐23 receptor and related T‐helper 17 cell cytokines in non‐small cell lung carcinoma
    Article Snippet: The commercial ELISA kits of IL‐23 and IL‐17 (R & D Systems) was carried out according to the manufacturer’s instructions. .. 2.11 Serum soluble IL‐23R bound with IL‐23 in serum of NSCLC by ELISA assay Interleukin‐23 recombinant protein (100 μL) (800 pg 1887‐ML‐010; R & D Systems) was coated in the ELISA plate at 4°C overnight, with assay buffer (1× PBS and 3% BSA) as negative control. .. After washing with washing buffer (0.05% Tween‐20) 3 times, 100 μL serum samples with high (50‐100 pg/mL) or low (20‐50 pg/mL) levels of sIL‐23R validated by ELISA from 20 NSCLC patients were incubated for 2 hours at room temperature.

    Negative Control:

    Article Title: Prognostic value of serum soluble interleukin‐23 receptor and related T‐helper 17 cell cytokines in non‐small cell lung carcinoma, et al. Prognostic value of serum soluble interleukin‐23 receptor and related T‐helper 17 cell cytokines in non‐small cell lung carcinoma
    Article Snippet: The commercial ELISA kits of IL‐23 and IL‐17 (R & D Systems) was carried out according to the manufacturer’s instructions. .. 2.11 Serum soluble IL‐23R bound with IL‐23 in serum of NSCLC by ELISA assay Interleukin‐23 recombinant protein (100 μL) (800 pg 1887‐ML‐010; R & D Systems) was coated in the ELISA plate at 4°C overnight, with assay buffer (1× PBS and 3% BSA) as negative control. .. After washing with washing buffer (0.05% Tween‐20) 3 times, 100 μL serum samples with high (50‐100 pg/mL) or low (20‐50 pg/mL) levels of sIL‐23R validated by ELISA from 20 NSCLC patients were incubated for 2 hours at room temperature.

    Cell Culture:

    Article Title: Immune requirements for protective Th17 recall responses to Mycobacterium tuberculosis challenge
    Article Snippet: Adoptive T cell transfer and experimental infections Naïve T cells were isolated from ESAT-6 Tg mice using CD4+ (L3T4) magnetic bead sorting (Miltenyi Biotec, San Diego, CA). .. To generate Th17 cells, CD4+ T cells were cultured at a 1:1 ratio with BMDCs in the presence of ESAT-61-20 peptide (10 μg/mL), recombinant (r)-mouse IL-2 (10 U/mL), r-mouse IL-6 (30 ng/ml R & D Systems), r-mouse IL-23 (50 ng/ml R & D Systems), r-human TGF-β (5 ng/mL R & D Systems), anti-IL-4 antibody (10 μg/mL), and anti-IFN-γ antibody (10 μg/mL) in Iscove’s Modified Dulbecco’s Medium (Life Technologies, Grand Island, NY). .. For Th1 cells, CD4+ T cells were cultured at a 1:1 ratio with BMDCs in the presence of ESAT-61-20 peptide (10 μg/mL), r-mouse IL-2 (10 U/mL), r-mouse IL-12 (10 ng/ml R & D Systems) and anti-IL-4 antibody (10 μg/mL) in Iscove’s Modified Dulbecco’s Medium (Life Technologies, Grand Island, NY).

    Article Title: Microbiota-derived butyrate suppresses group 3 innate lymphoid cells in terminal ileal Peyer’s patches
    Article Snippet: For intracellular cytokine staining of Foxp3, cells were fixed, permeabilized, and stained with FITC-conjugated anti-CD4 Ab, PE-conjugated anti-CD25 Ab, and APC-conjugated anti-Foxp3 Ab using a Foxp3 staining set (eBioscience, San Diego, CA), according to the manufacturer’s protocol. .. For intracellular cytokine staining of IL-22, PP cells were cultured with or without butyrate and then stimulated for 4 hr with PMA (50 ng/mL), ionomycin (500 ng/mL), and mouse recombinant IL-23 (40 ng/mL; R & D Systems, Minneapolis, MN). .. Brefeldin A (eBioscience) was added to the culture medium and cells were incubated for the final 1.5 hr.

    Modification:

    Article Title: Immune requirements for protective Th17 recall responses to Mycobacterium tuberculosis challenge
    Article Snippet: Adoptive T cell transfer and experimental infections Naïve T cells were isolated from ESAT-6 Tg mice using CD4+ (L3T4) magnetic bead sorting (Miltenyi Biotec, San Diego, CA). .. To generate Th17 cells, CD4+ T cells were cultured at a 1:1 ratio with BMDCs in the presence of ESAT-61-20 peptide (10 μg/mL), recombinant (r)-mouse IL-2 (10 U/mL), r-mouse IL-6 (30 ng/ml R & D Systems), r-mouse IL-23 (50 ng/ml R & D Systems), r-human TGF-β (5 ng/mL R & D Systems), anti-IL-4 antibody (10 μg/mL), and anti-IFN-γ antibody (10 μg/mL) in Iscove’s Modified Dulbecco’s Medium (Life Technologies, Grand Island, NY). .. For Th1 cells, CD4+ T cells were cultured at a 1:1 ratio with BMDCs in the presence of ESAT-61-20 peptide (10 μg/mL), r-mouse IL-2 (10 U/mL), r-mouse IL-12 (10 ng/ml R & D Systems) and anti-IL-4 antibody (10 μg/mL) in Iscove’s Modified Dulbecco’s Medium (Life Technologies, Grand Island, NY).

    Staining:

    Article Title: Microbiota-derived butyrate suppresses group 3 innate lymphoid cells in terminal ileal Peyer’s patches
    Article Snippet: For intracellular cytokine staining of Foxp3, cells were fixed, permeabilized, and stained with FITC-conjugated anti-CD4 Ab, PE-conjugated anti-CD25 Ab, and APC-conjugated anti-Foxp3 Ab using a Foxp3 staining set (eBioscience, San Diego, CA), according to the manufacturer’s protocol. .. For intracellular cytokine staining of IL-22, PP cells were cultured with or without butyrate and then stimulated for 4 hr with PMA (50 ng/mL), ionomycin (500 ng/mL), and mouse recombinant IL-23 (40 ng/mL; R & D Systems, Minneapolis, MN). .. Brefeldin A (eBioscience) was added to the culture medium and cells were incubated for the final 1.5 hr.

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    R&D Systems il 23
    IL-1β increases <t>IL-23</t> stimulated maintenance of the Th17 phenotype. A, Naïve T cells were activated, primed with TGF-β+IL-6+IL-1β for the first round and cultured for 2 additional rounds of stimulation in the presence of
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    IL-23 stimulates macrophages and IL-12 stimulates T cells to produce SP. Splenic T cells or macrophages isolated as described in the text were incubated for 6 h with or without rIL-12 or <t>rIL-23</t> at 0.5 ng/ml. SP was extracted from the cells after the 6 h of incubation and quantified by an ELISA. The data are means of three determinations from three separate experiments ± the standard deviation (SD). For cells versus cells plus IL-12 or cells plus IL-23, P
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    IRF4 expression levels were upregulated in dendritic cells from the IRF5-deficient mice. ( A ) Phosphorylated NF-κB p65 expression in DCs stimulated with PBS or 1 μg/mL R848 for a quarter of an hour or one hour was measured. ( B ) IRF4 mRNA expression levels in DCs before and after stimulation with 1 μg/mL R848 for 24 h were determined by quantitative RT-PCR. Data are obtained from duplicate samples from 12 mice in each group. Values are presented as the mean ± SEM of three independent experiments. ( C , E ) Representative flowcytometryplots of IRF4 + <t>IL-23</t> + DCs before ( C ) and after ( E ) stimulation with 1 μg/mL R848 for 3 h. ( D , F ) The frequency of IRF4 + IL-23 + DCs before ( D ) and after ( F ) stimulation with 1 μg/mL R848 for 3 h. Data are obtained from 5 mice in each group. Values are presented as mean the ± SEM of two independent experiments.
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    Flow cytometric analysis of the isolated cells. Cells at a concentration of 1 × 10 6 cells/ml were stained with <t>biotinylated</t> <t>anti-IL-23R,</t> avidin-FITC, and anti-CD4-PE according to the manufacturers' instructions (A, B, and C). For surface staining
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    IL-1β increases IL-23 stimulated maintenance of the Th17 phenotype. A, Naïve T cells were activated, primed with TGF-β+IL-6+IL-1β for the first round and cultured for 2 additional rounds of stimulation in the presence of

    Journal:

    Article Title: IL-23 promotes maintenance but not commitment to the Th17 lineage 1

    doi:

    Figure Lengend Snippet: IL-1β increases IL-23 stimulated maintenance of the Th17 phenotype. A, Naïve T cells were activated, primed with TGF-β+IL-6+IL-1β for the first round and cultured for 2 additional rounds of stimulation in the presence of

    Article Snippet: Since IL-23 was capable of maintaining the IL-17-secreting phenotype, it allowed us to determine if IL-23 mediated commitment to the Th17 lineage, commitment being defined as the ability of cells to maintain the IL-17-secreting phenotype in the presence of cytokines promoting the development of other subsets.

    Techniques: Cell Culture

    IL-23 maintains the IL-17 secreting phenotype without affecting cell expansion or survival. A, Naïve CD4+ T cells were activated, cultured in TGF-β+IL-6+IL-1β and blocking antibodies (anti-IFN-γ and anti-IL-4) for five

    Journal:

    Article Title: IL-23 promotes maintenance but not commitment to the Th17 lineage 1

    doi:

    Figure Lengend Snippet: IL-23 maintains the IL-17 secreting phenotype without affecting cell expansion or survival. A, Naïve CD4+ T cells were activated, cultured in TGF-β+IL-6+IL-1β and blocking antibodies (anti-IFN-γ and anti-IL-4) for five

    Article Snippet: Since IL-23 was capable of maintaining the IL-17-secreting phenotype, it allowed us to determine if IL-23 mediated commitment to the Th17 lineage, commitment being defined as the ability of cells to maintain the IL-17-secreting phenotype in the presence of cytokines promoting the development of other subsets.

    Techniques: Cell Culture, Blocking Assay

    IL-23 does not program commitment to the Th17 lineage. A, Naïve T cells were activated, primed with TGF-β+IL-6+IL-1β for the first round followed by 2 rounds of stimulation in IL-23 + IL-1β, were cultured for an additional

    Journal:

    Article Title: IL-23 promotes maintenance but not commitment to the Th17 lineage 1

    doi:

    Figure Lengend Snippet: IL-23 does not program commitment to the Th17 lineage. A, Naïve T cells were activated, primed with TGF-β+IL-6+IL-1β for the first round followed by 2 rounds of stimulation in IL-23 + IL-1β, were cultured for an additional

    Article Snippet: Since IL-23 was capable of maintaining the IL-17-secreting phenotype, it allowed us to determine if IL-23 mediated commitment to the Th17 lineage, commitment being defined as the ability of cells to maintain the IL-17-secreting phenotype in the presence of cytokines promoting the development of other subsets.

    Techniques: Cell Culture

    IL-1b increases responsiveness of the Il17 locus. A, Naïve T cells were activated, primed with TGF-β+IL-6+IL-1β for the first round and cultured for 2 additional rounds of stimulation with IL-23, IL-1β, or IL-23+IL-1β

    Journal:

    Article Title: IL-23 promotes maintenance but not commitment to the Th17 lineage 1

    doi:

    Figure Lengend Snippet: IL-1b increases responsiveness of the Il17 locus. A, Naïve T cells were activated, primed with TGF-β+IL-6+IL-1β for the first round and cultured for 2 additional rounds of stimulation with IL-23, IL-1β, or IL-23+IL-1β

    Article Snippet: Since IL-23 was capable of maintaining the IL-17-secreting phenotype, it allowed us to determine if IL-23 mediated commitment to the Th17 lineage, commitment being defined as the ability of cells to maintain the IL-17-secreting phenotype in the presence of cytokines promoting the development of other subsets.

    Techniques: Cell Culture

    IL-23 stimulates macrophages and IL-12 stimulates T cells to produce SP. Splenic T cells or macrophages isolated as described in the text were incubated for 6 h with or without rIL-12 or rIL-23 at 0.5 ng/ml. SP was extracted from the cells after the 6 h of incubation and quantified by an ELISA. The data are means of three determinations from three separate experiments ± the standard deviation (SD). For cells versus cells plus IL-12 or cells plus IL-23, P

    Journal: Infection and Immunity

    Article Title: Interleukin-12 (IL-12) and IL-23 Induction of Substance P Synthesis in Murine T Cells and Macrophages Is Subject to IL-10 and Transforming Growth Factor β Regulation

    doi: 10.1128/IAI.00358-08

    Figure Lengend Snippet: IL-23 stimulates macrophages and IL-12 stimulates T cells to produce SP. Splenic T cells or macrophages isolated as described in the text were incubated for 6 h with or without rIL-12 or rIL-23 at 0.5 ng/ml. SP was extracted from the cells after the 6 h of incubation and quantified by an ELISA. The data are means of three determinations from three separate experiments ± the standard deviation (SD). For cells versus cells plus IL-12 or cells plus IL-23, P

    Article Snippet: The isolated cells were cultured for 4 h in vitro with or without rIL-23.

    Techniques: Isolation, Incubation, Enzyme-linked Immunosorbent Assay, Standard Deviation

    IL-23 induces macrophages, while IL-12 induces T cells to express PPT A mRNA. Macrophages (F/480 + ) or T cells (Thy 1.2 + ) were positively selected from dispersed splenocytes of CBA/J schistosome-infected mice. Cells were cultured in the presence or absence of rIL-2 or rIL-23 at the indicated concentrations for 4 h. Cellular RNA was extracted, reverse transcribed, and subjected to quantitative RT-PCR to measure the PPT A transcript number after the incubation. The data are means of multiple determinations from three separate experiments ± the standard error (SE).

    Journal: Infection and Immunity

    Article Title: Interleukin-12 (IL-12) and IL-23 Induction of Substance P Synthesis in Murine T Cells and Macrophages Is Subject to IL-10 and Transforming Growth Factor β Regulation

    doi: 10.1128/IAI.00358-08

    Figure Lengend Snippet: IL-23 induces macrophages, while IL-12 induces T cells to express PPT A mRNA. Macrophages (F/480 + ) or T cells (Thy 1.2 + ) were positively selected from dispersed splenocytes of CBA/J schistosome-infected mice. Cells were cultured in the presence or absence of rIL-2 or rIL-23 at the indicated concentrations for 4 h. Cellular RNA was extracted, reverse transcribed, and subjected to quantitative RT-PCR to measure the PPT A transcript number after the incubation. The data are means of multiple determinations from three separate experiments ± the standard error (SE).

    Article Snippet: The isolated cells were cultured for 4 h in vitro with or without rIL-23.

    Techniques: Crocin Bleaching Assay, Infection, Mouse Assay, Cell Culture, Quantitative RT-PCR, Incubation

    PPT A expression in LP macrophages is subject to IL-23 and TGF-β regulation. Isolated LP macrophages from healthy mice were incubated 4 h with or without rIL-23 in the presence or absence of TGF-β all at 1 ng/ml. After the incubation, cellular RNA was extracted and analyzed for PPT A mRNA expression using PCR. The results from two individual experiments are shown. All samples contained comparable amounts of the HPRT housekeeping gene transcripts.

    Journal: Infection and Immunity

    Article Title: Interleukin-12 (IL-12) and IL-23 Induction of Substance P Synthesis in Murine T Cells and Macrophages Is Subject to IL-10 and Transforming Growth Factor β Regulation

    doi: 10.1128/IAI.00358-08

    Figure Lengend Snippet: PPT A expression in LP macrophages is subject to IL-23 and TGF-β regulation. Isolated LP macrophages from healthy mice were incubated 4 h with or without rIL-23 in the presence or absence of TGF-β all at 1 ng/ml. After the incubation, cellular RNA was extracted and analyzed for PPT A mRNA expression using PCR. The results from two individual experiments are shown. All samples contained comparable amounts of the HPRT housekeeping gene transcripts.

    Article Snippet: The isolated cells were cultured for 4 h in vitro with or without rIL-23.

    Techniques: Expressing, Isolation, Mouse Assay, Incubation, Polymerase Chain Reaction

    IRF4 expression levels were upregulated in dendritic cells from the IRF5-deficient mice. ( A ) Phosphorylated NF-κB p65 expression in DCs stimulated with PBS or 1 μg/mL R848 for a quarter of an hour or one hour was measured. ( B ) IRF4 mRNA expression levels in DCs before and after stimulation with 1 μg/mL R848 for 24 h were determined by quantitative RT-PCR. Data are obtained from duplicate samples from 12 mice in each group. Values are presented as the mean ± SEM of three independent experiments. ( C , E ) Representative flowcytometryplots of IRF4 + IL-23 + DCs before ( C ) and after ( E ) stimulation with 1 μg/mL R848 for 3 h. ( D , F ) The frequency of IRF4 + IL-23 + DCs before ( D ) and after ( F ) stimulation with 1 μg/mL R848 for 3 h. Data are obtained from 5 mice in each group. Values are presented as mean the ± SEM of two independent experiments.

    Journal: International Journal of Molecular Sciences

    Article Title: Exacerbated Imiquimod-Induced Psoriasis-Like Skin Inflammation in IRF5-Deficient Mice

    doi: 10.3390/ijms21103681

    Figure Lengend Snippet: IRF4 expression levels were upregulated in dendritic cells from the IRF5-deficient mice. ( A ) Phosphorylated NF-κB p65 expression in DCs stimulated with PBS or 1 μg/mL R848 for a quarter of an hour or one hour was measured. ( B ) IRF4 mRNA expression levels in DCs before and after stimulation with 1 μg/mL R848 for 24 h were determined by quantitative RT-PCR. Data are obtained from duplicate samples from 12 mice in each group. Values are presented as the mean ± SEM of three independent experiments. ( C , E ) Representative flowcytometryplots of IRF4 + IL-23 + DCs before ( C ) and after ( E ) stimulation with 1 μg/mL R848 for 3 h. ( D , F ) The frequency of IRF4 + IL-23 + DCs before ( D ) and after ( F ) stimulation with 1 μg/mL R848 for 3 h. Data are obtained from 5 mice in each group. Values are presented as mean the ± SEM of two independent experiments.

    Article Snippet: After surface staining, DCs were permeabilized using a fixation/permeabilization buffer (eBioscience) and PE-Cy7 anti-IRF4 antibody (BioLegend), PE anti-IL-23 antibody (R & D Systems), or isotype-matched control antibody (BioLegend) was added.

    Techniques: Expressing, Mouse Assay, Quantitative RT-PCR

    IRF5 deficiency downregulates IL-10 expression and upregulates IL-23 expression by dendritic cells. Dendritic cells (DCs) from the wild-type (WT) or IRF5 knockout (IRF5 KO) mice were stimulated with phosphate-buffered saline (PBS) or 1 µg/mL R848 for 24 h. Messenger RNA levels of IL-10 ( A ), IL-23p19 ( B ) and IL-12/23p40 ( C ) were determined by quantitative RT-PCR. Protein levels of IL-10 in the supernatants after R848 stimulation ( D ) IL-23 in the supernatants without R848 stimulation ( E ) were determined by enzyme-linked immunosorbent assay (ELISA). Data are obtained from duplicate samples from 12 mice in each group. Values are presented as mean the ± SEM of three independent experiments.

    Journal: International Journal of Molecular Sciences

    Article Title: Exacerbated Imiquimod-Induced Psoriasis-Like Skin Inflammation in IRF5-Deficient Mice

    doi: 10.3390/ijms21103681

    Figure Lengend Snippet: IRF5 deficiency downregulates IL-10 expression and upregulates IL-23 expression by dendritic cells. Dendritic cells (DCs) from the wild-type (WT) or IRF5 knockout (IRF5 KO) mice were stimulated with phosphate-buffered saline (PBS) or 1 µg/mL R848 for 24 h. Messenger RNA levels of IL-10 ( A ), IL-23p19 ( B ) and IL-12/23p40 ( C ) were determined by quantitative RT-PCR. Protein levels of IL-10 in the supernatants after R848 stimulation ( D ) IL-23 in the supernatants without R848 stimulation ( E ) were determined by enzyme-linked immunosorbent assay (ELISA). Data are obtained from duplicate samples from 12 mice in each group. Values are presented as mean the ± SEM of three independent experiments.

    Article Snippet: After surface staining, DCs were permeabilized using a fixation/permeabilization buffer (eBioscience) and PE-Cy7 anti-IRF4 antibody (BioLegend), PE anti-IL-23 antibody (R & D Systems), or isotype-matched control antibody (BioLegend) was added.

    Techniques: Expressing, Knock-Out, Mouse Assay, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

    Flow cytometric analysis of the isolated cells. Cells at a concentration of 1 × 10 6 cells/ml were stained with biotinylated anti-IL-23R, avidin-FITC, and anti-CD4-PE according to the manufacturers' instructions (A, B, and C). For surface staining

    Journal: Clinical and Vaccine Immunology : CVI

    Article Title: Interleukin-12 Is the Optimum Cytokine To Expand Human Th17 Cells In Vitro ▿

    doi: 10.1128/CVI.00022-09

    Figure Lengend Snippet: Flow cytometric analysis of the isolated cells. Cells at a concentration of 1 × 10 6 cells/ml were stained with biotinylated anti-IL-23R, avidin-FITC, and anti-CD4-PE according to the manufacturers' instructions (A, B, and C). For surface staining

    Article Snippet: The cells were stained with biotinylated anti-IL-23R (R & D Systems, Inc., Minneapolis, MN), avidin-fluorescein isothiocyanate (FITC) (eBioscience, San Diego, CA), and 20 μl of anti-CD4-phycoerythrin (PE) (eBioscience), according to the manufacturers' instructions.

    Techniques: Flow Cytometry, Isolation, Concentration Assay, Staining, Avidin-Biotin Assay